Home

User Guide - The Cambridge Crystallographic Data Centre

1. Weight 50 Run GOLD Run GOLD In The Background Finish Cancel e To highlight in the Hermes visualiser those protein atoms involved in a motif highlight the appropriate column and click on the View Atoms button To switch off highlighting click the Hide Atoms button e in order to remove all interaction and motif definitions click on Reset GOLD User Guide 11 Balancing Docking Accuracy and Speed 11 1 Num ber of Dockings GOLD will dock each ligand several times starting each time from a different random population of ligand orientations The results of the different docking runs are ranked by fitness score The number of dockings to be performed on each ligand is set when the ligand file is defined see Specifying the Ligand File s By default the number of dockings to be performed on each ligand is 10 The total time spent docking a ligand obviously depends on the number of docking runs so you can make GOLD go faster by reducing this number However it is useful to perform at least a few docking runs on each ligand This increases the chances of getting the right answer Also if the same answer is found in several different docking runs it is usually a strong indicator that the answer is correct The early termination option see Early Termination can be used to prevent GOLD wasting time performing multiple docking runs on easy ligands 11 2 Early Termination GOLD User Guide The earl
2. 1 1 1 1 Sorc OTICI 100 1 i i 1 i i i 1 1 1 1 1 i 1 it Tl Min ops l 10000 J Max ops 125000 Library Screening virtual Screening Ensemble Default Very Flexible Help R Run GOLD Run GOLD In The Background Finish Cancel e When using these ligand dependent GA settings the Search efficiency can be used to further control the speed of docking and the predictive accuracy i e the reliability of the results e With the Search efficiency set at 100 GOLD will attempt to apply optimal settings for each ligand For a ligand with five rotatable bonds this will be around 30 000 GA operations If the Search efficiency were set to 50 then GOLD will perform around 15 000 operations thereby speeding up the docking by a factor of two but the search space would be less well explored Similarly by setting a Search efficiency greater than 100 it is possible to make the search more exhaustive but slower e The following search efficiency settings are available by clicking the corresponding button Very flexible this sets the search efficiency at 200 and is recommended for large highly flexible ligands This setting delivers high predictive accuracy but is relatively slow Default this sets the search efficiency at 100 GOLD will attempt to apply optimal settings for each ligand see above Virtual screening this sets the search efficiency at 30 this setting is suitable for rout
3. To fix all rotatable bonds in the ligand at their input conformation select the fix all button To fix all non terminal rotatable bonds i e not CH3 OH etc select the fix all but terminal button To fix the rotatable bond between two specified atoms select fix specific then click on the Specify Bonds button The resulting Select Ligand Bonds To Fix dialogue allows you to select the bond s you wish to fix To select a bond hit Add then either select the bond by clicking on it in the visualiser or by entering the bond atom indices directly Multiple rotatable bonds can be specified Click on Delete to remove a bond from the list Once you are satisfied with your selections click on Close The ability to fix rotatable bonds in the ligand at their input conformations is also available using the rotatable _ bond _ override mol2 file see Overriding Automatic Bond Settings This is particularly useful if docking a library of ligands that have a common substructure rather than the method above which is more suitable when docking an individual ligand Note When fixing all rotatable bonds at their input conformation i e performing a rigid ligand docking GOLD will try to find the best orientation of the ligand in the binding site by mapping donor acceptor as well as hydrophobic hydrophobic fitting GOLD User Guide points However GOLD will not perform a local optimisation simplex on the final solution This may lead to pe
4. Constraints Distance HBond Covalent Help R Protonation amp Tautomers Delete Ligands Flexible Sidechains Constraint weight 10 0 Minimum H bond geometry weight 0 005 Soft Potentials Substructure Protein HBond Interaction Motif Protein atomis required to form H bond 460 Protein HBond 10 0 0 005 1162 1161 Protein HBond 10 0 0 005 1380 Protein HBond 10 0 0 005 241 242 Protein HBond 10 0 0 005 460 Delete Clear Run GOLD Run GOLD In The Background Finish Cancel Running GOLD GOLD User Guide Return to the general docking setup by clicking on the Global Options tab Click on Output Options Either type an output directory name in the Output directory window or browse to a directory using the button adjacent to this window This is where the GOLD output files will be written Click on the Run GOLD button at the bottom of the GOLD front end All settings can remain as they are so hit Save to start the GOLD run You will be prompted that a file called gold conf already exists and asked if you want to overwrite it Click OK to agree to overwrite the existing gold conf file Alternatively choose Cancel to go back to the Finish GOLD Configuration window enter a new file name for the GOLD conf file and press Save The GOLD job will now start interactively As the job progresses output will be displayed in the Run GOLD window The Run GOLD output window
5. To dock into a rigid protein select Proteins from the list of Global Options given on the left of the GOLD Setup window and activate the Fix all protein rotatable bonds tick box 3 9 Metal lons 3 9 1 Preparing a Protein Input File which Contains a Metal lon There are some additional requirements when preparing a protein input file which contains a metal ion The metal ion must be coordinated to at least two protein atoms or water molecules so that GOLD can predict the coordination geometry see Automatic Determination of Metal Coordination Geometries In the protein input file the metal ion should not have any bonds to coordinating atoms If these are present in the original PDB file they must be deleted Note GOLD can only handle the hardcoded metal atom types see Automatic Determination of Metal Coordination Geometries it is not possible to add user defined metal atom types If a particular metal ion is not required it can be removed from the protein see Deleting Ligands and Metal lons 3 9 2 Automatic Determination of Metal Coordination Geometries 20 GOLD is able to recognise the following metal coordination geometries Template Geometry Coordination Number TETR Tetrahedral n 4 GOLD User Guide Template Geometry Coordination Number TBP Trigonal bipyramidal n 5 OCT Octahedral n 6 CTP Capped trigonal prism n 7 PBP Pentagonal bipyramidal n 7 SQAP Square prism n 8 ICO Icosahedral n 10 DOD Dodecahedral n
6. Constraint contribution to Chemscore value Contribution for weak CH O H bonds Internal ligand energy offset Protein energy term to penalise clashes when using flexible sidechains Penalty term for non displacement of active site waters RMSd of solution against reference ligand Total ASP fitness value of docked ligand Calculated statistical potential plus the ChemScore clash term and internal energy term The total calculated statistical potential is a summation over all combinations of protein and ligand atoms Protein ligand H bond contribution to ASP value Metal binding contribution to ASP value See see Metal Binding and Lipophilic Terms see Metal Binding and Lipophilic Terms see Hydrogen Bond Terms see Clash Penalty and Internal Torsion Terms see Clash Penalty and Internal Torsion Terms see Overview see Covalent Term see Constraint Terms see Kinase Scoring Function see Internal Energy Offset see Protein Protein Clashes see Water Molecules see Specifying a Ligand Reference File see Astex Statistical Potential ASP see Astex Statistical Potential ASP see Astex Statistical Potential ASP see Astex Statistical Potential ASP see Astex Statistical Potential ASP 223 Name Gold ASP DEClash Gold ASP DEInternal Gold ASP Rot Gold ASP Covalent Gold ASP Constraint Gold ASP Protein Energy Gold ASP SBar
7. GoldMine Parallel GOLD Save ligand log files Constraints Atom Typing IV Save initialised ligand files I Save solutions to one file es J Use alternative bestranking Ist filename Vy J Create links for different binding modes based on RMSD clustering Distance between clusters Jo 75 A Help R Run GOLD Run GOLD In The Background Finish Cancel e Each ligand will normally be docked several times so a given input ligand will produce a set of files each containing the results of a separate docking attempt Alternatively you can specify that all saved docking solutions for all ligands are to be concatenated and written to a single file To do this enable to Save solutions to one file check box and either enter the path and filename of the file or click on the button and use the file selection window to choose the file 118 GOLD User Guide 13 2 Controlling the Information Written to Ligand Solution Files It is possible to write additional information to docked solution files This information is written to SD file tags for MOL2 files these tags are written to comment blocks For post processing docking results with GoldMine it is particularly important that the scoring function terms and the rotated protein positions are saved Click on Output Options from the list of Global Options given on the left of the GOLD Setup window then select the Information in File tab GOLD Setup
8. HBOND_COEFFICIENT see ChemScore METAL_COEFFICIENT see ChemScore CHARGED_HBOND_FACTOR Scaling factor for charged hydrogen bonds expected to be greater than or equal to one CHARGED_METAL_FACTOR Scaling factor for charged acceptors coordinating to a metal ion expected to be greater than or equal to one DELTA_BETA_IDEAL see ChemScore DELTA_BETA_MAX see ChemScore CHO_COEFFICIENT see ChemScore CHO_TYPE see ChemScore CHO_R_IDEAL see ChemScore CHO_DELTA_R_IDEAL see ChemScore CHO_DELTA_R_MAX see ChemScore CHO_ALPHA_IDEAL see ChemScore CHO_DELTA_ALPHA_IDEAL see ChemScore CHO_DELTA_ALPHA_MAX see ChemScore CHO_BETA_IDEAL see ChemScore CHO_DELTA_BETA_IDEAL see ChemScore CHO_DELTA_BETA_MAX see ChemScore HBOND_SCALING see ChemScore lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt string gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt GOLD User Guide 7 3 GoldScore 7 3 1 Overview GOLD User Guide The GOLD fitness function is made up of four components protein ligand hydrogen bond energy external H bond protein ligand van der Waals vdw energy external vdw
9. ligand internal vdw energy internal vdw ligand torsional strain energy internal torsion Optionally a fifth component ligand intramolecular hydrogen bond energy internal H bond may be added If any constraints have been specified then an additional constraint scoring contribution S con will be made to the final fitness score Similarly when docking covalently bound ligands a covalent term S cov will be present By default output files will contain a single internal energy term S int which is the sum of the internal torsion and internal vdw terms To write these component terms to output files you will need to edit the gold params file see Altering GOLD Parameters the gold params File to include the following line VERBOSE_SCORE 1 Empirical parameters used in the fitness function hydrogen bond energies atom radii and polarisabilities torsion potentials hydrogen bond directionalities etc are taken from the GOLD parameter file These parameters are independent of the scoring function being used Parameters can be customised by copying the file editing the copy and instructing GOLD to use the edited file see Altering GOLD Parameters the gold params File A scoring function specific parameters file is also used for GoldScore this is called goldscore params Parameters within this file can also be modified see Altering GoldScore Fitness Function Parameters the GoldScore Parameters File Th
10. From within the 1TBF tab add hydrogen atoms to the protein by selecting the Add Hydrogens button from the first Protonation amp Tautomers option in the Wizard Note that all hydrogen atoms must be present in the protein file prior to docking The hydrogen atoms are placed on the protein in order to ensure that ionisation and tautomeric states are defined unambiguously When adding hydrogen atoms you can protonate using the standard GOLD protonation rules or you can use user defined SMARTS based rules by enhancing the protonation rules text file supplied in the GOLD distribution Still in the 1TBF tab hit the Extract Delete Waters option in the Wizard From within this dialogue it is possible to specify water molecules that mediate protein ligand interactions i e active waters and to delete those that are not required Since you do not want to extract any waters for docking hit the Delete Remaining Waters button When prompted Are you sure you want to delete all the waters hit OK You will be informed that 323 waters have been deleted 211 Return to the Global Options tab of the GOLD Wizard To superimpose 1tbf with the other three proteins hit the Superimpose Proteins button then follow the onscreen instructions You can use a component of the FASTA package or if binaries can t be found the default is to use Needleman Wunsch algorithm Both FASTA and the Needleman Wunsch algorithm do the same thing i e they generate global sequen
11. Gold ASP Internal Correction Gold ASP Reference RMSD Piecewise Linear Potential PLP Gold PLP Fitness Gold PLP PLP Gold PLP part hbond Gold PLP part metal Gold PLP part buried Gold PLP part nonpolar Gold PLP part repulsive Gold PLP ligand clash Gold PLP ligand torsion Gold PLP Chemscore hbond Gold PLP Chemscore CHOscore 224 Explanation Protein ligand clash penalty to the ASP value Internal ligand intramolecular H bond contribution to ASP value Rotatable bond freezing term contribution to ASP value Covalent bonding contribution to ASP value Constraint contribution to ASP value Protein energy term to penalise clashes when using flexible sidechains Penalty term for non displacement of active site waters Internal ligand energy offset RMSd of solution against reference ligand Total PLP fitness value of docked ligand Calculated potentials plus the ChemScore clash term and internal energy term Protein ligand H bond contribution to PLP value Metal binding contribution to PLP value Scoring contribution from buried interaction types Scoring contribution from nonpolar interaction types Scoring contribution from repulsive interaction types Protein ligand clash penalty to the PLP value Internal ligand torsional strain penalty to the PLP value Chemscore Protein ligand H bond contribution Contribution for weak CH O H bonds See see Astex Statistica
12. S 02 0 c02 ar O co2 Sulphone 0 2 0 2 ue oc e Sulfoxide sulfinyl 0 2 S o N R NR 6 5 Internal GOLD Atom Types e GOLD uses four internal atom types which are not recognised by SYBYL These are N plc nitrogen donors in a protonated delocalised system such as a guanidinium ion N acid acidic nitrogen e g in tetrazole or sulphonamide ions S a sulphur acceptors and S m charged sulphur atoms You should not really need to know about these but all assignments of the N plc N acid S a and S m atom types are logged in the gold log file so you can check to see if everything is working as you would expect GOLD User Guide 53 7 7 1 7 2 7 2 1 54 Fitness Functions Selecting a Fitness Function GOLD offers a choice of fitness functions GoldScore see GoldScore ChemScore see ChemScore ASP see Astex Statistical Potential ASP CHEMPLP see Piecewise Linear Potential CHEMPLP and User Defined Score see User Defined Scoring Function CHEMPLP has been found to give the highest success rates for both pose prediction and virtual screening experiments against diverse validation test sets and is therefore the default scoring function in GOLD With respect to use of the GoldScore ChemScore ASP they are about equally reliable although on any given problem one may give a good prediction and the other not Therefore when screening large numbers of compounds rescoring docking poses with alternative
13. Setting Up Substructure Based Distance Constraints To use a substructure based distance constraint first create a file containing the substructure in mo12 format e g substructure mol2 The actual conformation of the group in this file is not important as only the element types and 2D connectivity will be used To set up a distance constraint you must first select the appropriate protein tab adjacent to the Global Options tab To constrain a distance click on Distance from the list of Global Options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options To specify the Substructure file either enter the path and filename of the file or click on the Substructure File button and use the file selection window to choose the file Specify the Protein atom number and Substructure atom number be used in the distance constraint This can be done by clicking on an atom in the visualiser Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box Specify the allowed range of separation by entering a Maximum separation and a Minimum separation distances are in A Enter the spring constant i e the weight of the term This causes a spring based distance constraint to be added for the specified substructure atom and protein atom The weight specifies the spring energy
14. Bis Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Help R Wizard Templates Proteins s v Define Binding Site IV Save fitness score Select Ligands IV Weighted terms Waters Ligand Flexibility JV Unweighted terms Fitness amp Search Options GA Settings T Do not write SD style tags to Mol2 files ee Preserve COMMENT fields from input Mol2 ligand files Parallel GOLD Constraints Atom Typing IV Save lone pairs File Format Options Information in File Selecting Solutions IV Save protein rotated atom positions IV Save per atom scores Save per atom scores to charge field Run GOLD Run GOLD In The Background Finish Cancel GOLD User Guide The following options are available Save fitness score Enable this check box if you want the docked solution files to include the docking score terms e g the total GoldScore fitness value for each docking and its components such as protein ligand H bond energy internal ligand strain energy etc Weighted terms Certain docking scoring function terms are the product of a term dependent on the magnitude of a particular physical contribution e g hydrogen bonding and a scale factor determined e g by a regression coefficient The docking scoring function terms included in the output file can therefore consist of weighted terms non weighted
15. Changes to genetic algorithm parameters should be made with care see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 23 6 Operator Weights Migrate Mutate Crossover GOLD User Guide The operator weights are the parameters Mutate Migrate and Crossover or pt_cross They govern the relative frequencies of the three types of operations that can occur during a genetic optimisation point mutation of the chromosome migration of a population member from one island to another and crossover sexual mating of two chromosomes Each time the genetic algorithm selects an operator it does so at random Any bias in this choice is determined by the operator weights For example if Mutate is 40 and Crossover is 10 then on average four mutations will be applied for every crossover The migrate weight should be zero if there is only one island otherwise migration should occur about 5 of the time Changes to genetic algorithm parameters should be made with care see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 227 24 24 1 228 Appendix E Utility Programs smart_rms rms_analysis check_mol2 are located in C Program Files x86 CCDC goldsuite 5 2 GOLD gold d_win32 bin and gold_utils is located in C Program Files x86 CCDC goldsuite 5 2 Hermes gold_utils exe smart_rms Located in C Program Files x86 CCDC goldsuite 5 3 GOLD gold d_win32 bin on Windows ma
16. Cluster 3 bestranking structure is gold_soln ligand m1 _ 4 mol2 Cluster 4 bestranking structure is gold_soln ligand _ml_ 9 mol2 GOLD can be set up to generate diverse solutions based on cluster size and RMSD see Generating Diverse Solutions Viewing Docked Solutions in Hermes Once the job is complete to visualise docked solutions in the Hermes visualiser click on the View Solutions button in the Run GOLD window Within Hermes the docking poses from GOLD docking jobs can be navigated and visualised alongside the associated protein model using the Docking Solutions pane of the Molecule Explorer situated to the left of the Visualiser window Numerical data associated with the solutions such as the fitness score and its components are tabulated as columns within the Docking Solutions pane The data can be sorted and or colour coded according to selected data columns Poses can be grouped Poses may also be manually selected and can then be re exported with a tailorable number of fields of associated data It is also possible in Hermes to further describe the docking poses by calculating additional descriptors for them These descriptors can be added to a GoldMine DB and used in further analysis The descriptors quantify amongst other things the hydrogen bonding interactions that occur between protein and docked ligand H bond interactions that do not occur e g a protein H bond donor that is prevented from forming a hydrogen
17. FIM SQLite Filename 7 Browse P Postgres aaa Database R gt nweJ2J2 nnnnn0 Hast PRE User R i Password TT Dock set Read dock sets Help R Run GOLD Run GOLD In The Background Finish Cancel e Click on the Run GOLD button GOLD will start by initialising the protein The GOLD server is now activated and ready to receive ligands for docking GOLD User Guide 43 e Next within GoldMine Open a GoldMine DB and make a selection of docking poses of interest within the Selection Manager window At the top right of the Selection Manager is an option marked Dock in GOLD Click on this button e Adialog window will appear informing you that a GOLD process set up to receive ligands from GoldMine is required Hit OK A GOLD dockings window is now displayed GOLD dockings 2 Eq Queued selections Connected processes ash ccde cam ac uk LIG_THRO_31_2 Received solutions Dock set 412 e The top segment displays the number of ligands queued for docking and the middle segment informs which ligand is currently being docked The docked run is complete once the word Waiting appears alongside the host name in the middle segment e Asecond selection can created and then submitted to the same GOLD process The results will be treated as though they are contiguous with the first set
18. GLN192 window Collectively these lines define the torsional flexibility that the Gln192 side chain will be allowed to have during docking 190 There are a number of parameters listed alongside each rotamer Chil is the first rotatable torsion in the side chain In the case of GLN192 this corresponds to rotation around Ca CB so the atoms will be the backbone N atom 2817 CA 2818 CB 2821 and CG 2822 Chi2 is the second rotatable torsion and corresponds to rotation around CB Cy so the atoms are CA 2818 CB 2821 CG 2822 and CD 2823 Chi3 is the third rotatable torsion corresponding to rotation around Cy Cd so the atoms are CB 2821 CG 2822 CD 2823 and terminal N 2825 Thus Rotamer1 specifies the first set of allowed values for chi1 chi2 and chi3 i e Chil 62 Chi2 180 Chi3 20 Associated with each numbered chi value is a delta value For Rotamer1 these are Delta1 13 Delta2 14 and Delta3 16 These delta values specify the allowed range e g delta1 chi1 to delta1 chi1 Each rotamer therefore describes one allowed conformation of the side chain as defined by the torsion angles values chi1 chi2 chi3 etc and their allowed ranges delta1 delta2 delta3etc GOLD User Guide e The dials at the top of the window reflect the rotamer information for the currently loaded rotamer In the first instance the dials reflect the settings for Rotamer1 Allowed rotation v
19. Ligand Score Contributions Protein Score Contributions GOLD User Guide Explanation List of protein residues used to define the binding site Optimised positions of polar protein hydrogen atoms that are generated during docking Optimised positions of water hydrogen atoms generated during docking Optimised torsions for rotatable bonds in the ligand Also for protein side chain torsions which have been specified as being allowed to rotate during docking Enabling the association of a solution with its protein Ligand identifier Rmsd of rescored solutions Total fitness value of docked ligand Covalent bonding contribution to the fitness score Constraint contribution to the fitness score For individual ligand atom it s scoring contribution to the total fitness score and also the constituent scoring terms For individual protein atom its scoring contribution to the total fitness score and also the constituent scoring terms See see Defining a Binding Site from a List of Atoms or Residues see File Containing the Protein Binding Site Geometry see Water Molecules see Side Chain Flexibility see Rescoring see Controlling the Information Written to Ligand Solution Files see Controlling the Information Written to Ligand Solution Files 221 Name Gold Ensemble ID GoldScore Gold Goldscore Fitness Gold Goldscore External Hbond Gold Goldscore External Vdw Gold Go
20. but they could be read in at run time if required e Directives are allowed to take account of special circumstances There are two directives expand and period e The expand directive has the form expand lt min gt lt max gt where lt max gt lt min gt 180 0 or lt min gt 0 This directive is used for torsions where the CSD query has GOLD User Guide symmetry and torsions are only measured over lt min gt to lt max gt degrees However although the CSD query may have two fold symmetry often the matched structure does not The expand directive fills out the rest of the histogram with the correct values e The period directive takes account of those torsional distributions for which the matched structure has symmetry This directive has the form period lt pmin gt lt pmax gt The distribution will only be expanded between angles lt pmin gt and lt pmax gt 25 3 Example Torsion Angle Distributions Here are some examples of torsion angle distributions extracted from the Cambridge Structural Database and in the correct format DIAGRAM acid T1 C 2 O co2 O co2 C 3 2H C 3 2H C 41800000001872000011000104101000 002241 DIAGRAM acid T2 O co2 C 2 O co2 C 3 2H C 3 2H C 8513213232334032711159141023144 1 3 3 60357 DIAGRAM amide nh T2 C 2 O 2 N am 1H C 3 1H C 3 N am 1H C 2 0 2 1 114 16 29 25 23 38 35 50 8215653 6 1000000 1 114171544 21252200 DIAGRAM uracil O
21. gt lt Gold Protein ActiveResidues gt The list must end with a blank line or the end of the text file GOLD will read multiple residue names from one line but lines must not exceed 250 characters in length Residue names must be separated by a space for example gt lt Gold Protein ActiveResidues gt HIS69 ARG71 GLU72 ARG127 ASN144 ARG145 GLY155 ALA156 GLU163 THR164 HIS196 SER197 TYR198 SER199 LEU201 LEU203 ILE243 ILE244 ILE247 TYR248 GLN249 ALA250 GLY253 SER254 ILE255 THR268 GLU270 PHE279 ZN309 All solvent accessible protein acceptor and donor atoms available to the ligand are taken from the list The file should contain all atoms or residues which are required to explicitly define the protein active site Click on the View button to highlight in the Hermes visualiser those residues that have at least one of their atoms included in the binding site definition The cavity atom selection can be saved as a protein atom subset and viewed within Hermes To do this click on the Add Definition as a Selection button You can then highlight the atoms belonging to the subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area Cavity Detection The binding site can be defined in several ways e g by specifying the approximate centre of the binding site and taking all atoms that lie within a specified radius of this point see Defining a Binding Site from
22. hydrophobic constraint calculation including the final contribution to the fitness score are given in the ligand log file see Ligand Log File 10 4 2 Setting Up Region Hydrophobic Constraints e To define a region hydrophobic constraint click on Region from the list of Global Options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options e Specify the ligand atoms to be used in the constraint by selected either All hydrophobic atoms Hydrophobic atoms in aromatic rings or User specified list If User specified list is selected then individual ligand atoms can be selected by clicking on them in the visualiser you will need to first hide the sphere using the Centroid visible checkbox in the Edit Sphere dialogue Alternatively you can enter the atom numbers as it appears in the input file directly into entry box e Next specify the position and radius of the sphere To do this click on the Define Sphere button this will launch the Edit Sphere dialogue 98 GOLD User Guide gt GOLD Setup yx Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial gold conf Load Save Options Protein 1cil lyasefoxo acid Wizard Templates Ligand atom selection Proteins Define Binding Site all hydrophobic atoms Select Ligands Waters Hydrophobic atoms in aromatic rings Ligand Flexi
23. see ChemScore Astex Statistical Potential ASP see Astex Statistical Potential ASP and User Defined Score see User Defined Scoring Function Ensure that the default CHEMPLP scoring function is selected A number of additional options are available by clicking on the More gt gt button Allow early termination by default the Allow early termination check box should be switched on Click on the Early Termination Options button to inspect the settings Early Termination Options 21 x Terminate the number of GA runs early ifthe top f solutions are within 1 5 A Close This will instruct GOLD to terminate the docking if at any point the best three solutions found are all within 1 5 A rmsd of each other In this case it 149 is probable that the answer is correct and further docking runs will not be required Keep the settings as they are and hit Close For the purposes of this tutorial all other settings should be left at their default values Hit the Next button to proceed to the GA Search Options window Selecting Docking Speed GOLD optimises the fitness score using a genetic algorithm GA see Genetic Algorithm Overview A number of parameters control the precise operation of the genetic algorithm The settings are encapsulated into three speeds Slow most accurate this equates to 100 000 operations Medium 50 000 operations Fast least accurate 10 000 operations Further opt
24. see Hydrogen Bond Constraints To apply one of the above constraints it is necessary to first click on the appropriate protein tab e g pdb1qpc_full_entry below adjacent to the Global Options tab To define a constraint select a constraint type from those listed on the left of the GOLD Setup window If individual constraint types are not visible click on the icon next to Constraints to expand the list of options 89 3 GOLD Setup BBE Conf file C Documents and Settings henderson Desktop ensemble files ensemble conf Load Save Options pdbigpc_full_entry pdbigpd_full_entry pdbigpe_full_entry pdb1gpj_full_entry pdb3ick_a Protonation amp Tautomers Waters Delete Ligands Flexible Sidechains Soft Potentials Metals SE Constraints The following constraints are available for definition here Distance Substructure HBond Protein HBond Covalent Interaction Motif Distance constrain the distance between specific atoms e Substructure distance constraints for use with multiple ligands HBond favour Formation of a specified H bond Protein HBond bias certain protein atoms to form H bonds Delete Clear Help h Run GOLD Run GOLD In The Background Finish Cancel e Constraints that are applicable to an individual protein or protein ensemble are the following Region hydrophobic constraint for biasing the docking towards solutions in which particu
25. select atoms to define a centroid or edit XYZ Ligand Flexibility Fitness amp Search Options GA Settings x 515 y 1 76885 Z 14 2075 View Reset Output Options GoldMine Parallel GOLD One or more ligands i E Constraints Atom Typing List of atoms or residues Filename a View Select all atoms within 10 amp J Generate a cavity atoms file From the selection Refine Selection IV Detect cavity restrict atom selection to solvent accessible surface J Force all H bond donors acceptors to be treated as solvent accessible Add Definition as a Selection Help R Run GOLD Run GOLD In The Background Finish Cancel e The approximate radius of the binding site must also be specified If r is the radius the binding site will be defined as all atoms that lie within r of the specified protein atom By default the binding site radius is set to 10 0 This can be changed by entering a value in the box labelled Select all atoms within e Residues that have at least one of their atoms included in the binding site definition will be highlighted in the Hermes visualiser When entering a new value in the Select all atoms within box it is necessary to hit the enter key before the visualiser will update to reflect the changes made GOLD User Guide 13 3 6 3 14 After visual inspection you may wish to manually refine the binding site definition To do this switch on
26. shown coloured in green via interaction with the sulphonamide N atom Metal coordination in GOLD is modelled as pseudo hydrogen bonding Metal ligand interactions will typically involve the metal binding to for example carboxylate ions deprotonated histidines i e negatively charged and phenolates Therefore metals can be considered to bind to H bond acceptors and the metal will compete with H bond donors for interaction GOLD User Guide This ends the tutorial GOLD User Guide 163 20 3 Tutorial 3 Use of Hydrogen Bonding Constraints Introduction Input Files 164 First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial3 toa directory to which you have write permissions The design of new and more potent antiretroviral agents for the human immunodeficiency virus HIV continues to be the focus of much attention The crystal structures of HIV 1 protease in complex with a number of cyclic urea inhibitors have been determined in order to identify the key interactions responsible for the high potency of this class of inhibitor see Jadhav et al J Med Chem 40 181 1997 The C symmetric cyclic urea scaffold is well suited to interact with the viral protease It has been observed that these inhibitors are anchored in the active site of the protease by six key hydrogen bonds The object of this tutorial is to investigate the binding mode of a cyclic urea inhibitor with HIV 1 protease PDB
27. the v terms are the regression coefficients and the P terms represent the various types of physical contributions to binding The final ChemScore value is obtained by adding in a clash penalty and internal torsion terms which militate against close contacts in docking and poor internal conformations Covalent and constraint scores may also be included j c binding clash internal internal covalent covalent constraint Empirical parameters used in the fitness function hydrogen bond energies atom radii and polarisabilities torsion potentials hydrogen bond directionalities etc are taken from the GOLD parameter file These parameters are independent of the scoring function being used Parameters can be customised by copying the file editing the copy and instructing GOLD to use the edited file see Altering GOLD Parameters the gold params File A scoring function specific parameters file is also used for ChemScore this is called chemscore params Parameters within this file can also be modified see Altering ChemScore Fitness Function Parameters the ChemScore File Block Functions in ChemScore ChemScore uses block functions throughout its implementation to describe contact terms of various types A block function is of the following form 61 lif x lt Da Blax x 410 _ 2 uai ifs lt x lt x ideal max z 7 ideal max A A max ideal Oif x gt x max e This functional form looks like X ide
28. 1 9 Ligand Log File e The progress of each genetic algorithm run is listed in the ligand log file gold lt ligand_ file name gt _m 1log Here m is an index to the number of the ligand in the input file e g m3 indicates that the log file refers to the third ligand in the input ligand file remember that an input file may contain more than one ligand e The log files are line buffered so you can see how the algorithm is progressing even when GOLD is run in the background e The parallel version of GOLD creates several temporary log files for each ligand named gold_soln lt ligand_ file name gt _m _ lt N gt 1log where lt N gt is a docking run number Once all the docking runs for the ligand have been completed these files are concatenated together into the single log file gold soln lt ligand_file name gt m log e The ligand log file contains information on The progress of each docking run see Information on the Progress of Docking Runs A comparison of the various docking solutions found see Comparison of Docking Solutions Clustering of ligand poses for identification of solutions with different binding modes see Identification of Different Binding Modes Clustering of Ligand Poses e If you do not wish to save ligand rank files click on Output Options from the list of Global Options given on the left of the GOLD Setup window select the File Format Options tab then disable the Save ligand log files
29. 1 has the worst fitness GOLD User Guide Final ranked order of GA solutions 2 RMSD Matrix of RANKED solutions 2 3 Lot 1 0 I3 2 3 0 7 Clustering method complete linkage Structure ids in cluster table rank nos Ordering of clusters and their members by rank Corder if from rms_analysis Distance Clusters O 75 1 2 3 1 26 ae se Finished Docking Ligand C Program Files CCcDC GOLD examp les e We have finished with the Run GOLD window now so close the window by clicking on the Close button Fitness Function Rankings Files ligand_m1 rnk and bestranking Ist e The ligand_m1 rnk file is stored in the specified output directory open and inspect the file in a text editor This file contains a summary of the fitness scores for all the docking attempts on the N phosphonacetyl L aspartate ligand e The docking attempts are listed according to fitness score so the best solution is placed first e The file gives total fitness scores and a breakdown of the fitness into its constituent energy terms e 6A file called bestranking st is written and gives a continuous summary of the best solution that has been obtained for each completed ligand The file gives total fitness scores and a breakdown of the fitness into its constituent energy terms Files Containing the Docked Ligand gold_so n_ligand_m _n mol2 e The N phosphonacetyl L aspartate ligand will have been docked a number of times so a set of files will have b
30. 1ACJ soln 1 37 7872 tacrine ligand from 1ACJ soln 2 37 8299 tacrine ligand from 1ACJ saln 3 37 6039 tacrine ligand from 1ACJ saln 4 37 9719 tacrine ligand from 1ACJ soln 5 37 8133 tacrine ligand from 1ACJ soln 6 37 6595 tacrine ligand fram 1ACJ saln 37 6584 tacrine ligand from 1ACJ saln 8 37 9411 tacrine ligand from 1ACJ saln 9 37 9441 i facrine igand from 1ACJ soln 10 37 8066 The solutions are numbered sequentially thus enabling you to establish which docking run they correspond to The reference ligand is still displayed Check each solution in turn against the pose of the reference ligand Now it is likely that only one docking mode is represented This docking mode is close to that of the reference ligand It is not a perfect superposition though as the ligand attempts to contact the protein along its edge more closely than it does in reality The values of the docking scores for this run are higher than those of the previous run Analysis of results All waters toggled GOLD User Guide From Load GOLD run results in Hermes read in the gold conf file corresponding to the waters_toggle GOLD run The reference ligand is still displayed so compare its pose to the solutions you ve just loaded The top ranking pose is now much closer to that of the reference ligand Also the scores for this run are higher than the two previous runs Notice that the two waters able to interact with the NH of the ligand have also b
31. 4 7 el 6 910 1 17 Lo 3 405 fF 8 1 6 93 10 6 36 12 3 4 5 6 7 8 910 Links have been produced for each cluster Cluster 1 bestranking structure is gold_soln_ligand_m1_8 mo12 Cluster 2 bestranking structure is gold_soln_ligand_m1_10 mo12 Cluster 3 bestranking structure is gold_soln_ligand_m1_4 mol12 Cluster 4 bestranking structure is gold_soln_ligand_m1_9 mol12 e Inthe above example at a clustering distance of 0 75 A there are four different clusters of solutions 0 90 1235 47 6910 8 files d 0 75 A Clusters are separated by the symbol and rankings are used rather than run numbers see Files Containing the Docked Ligand s e The first cluster contains four solutions ranked numbers 1 2 3 and 5 the bestranking structure in this cluster is ranked_structure_m _1 mol2 which corresponds to the docked solution gold_soln_ ligand_ml_8 mol12 Likewise the second cluster contains two solutions ranked numbers 4 and 7 the bestranking structure in this cluster is ranked_structure_m _4 mol2 which corresponds to the docked solution gold_soln_ligand_m1_10 mo12 and so on for the fourth and fifth clusters GOLD User Guide 133 15 5 15 6 15 6 1 134 Symbolic links will be generated in the output directory which will link to the top ranked solution in each cluster Cluster 1 bestranking structure is gold_soln ligand_m1_ 8 mol Cluster 2 bestranking structure is gold_soln ligand _ml_10 mol2
32. 7 3 3 4 Specifying Histidine Tautomers csseeeseecceceecsaeeeseeeeeeeeeseaaeaseeeeeess 7 3 4 Deleting Ligands and Metal TONS esineisiin ieir eiia 8 3 5 Water Molecules ccc cccccceeeeeeencceeeeeeeeeeeeenaaeeeeeeeeeeeeeaaaaeeeeeeeeeeeaaaaeeeeeeeeeeeeaaaaes 9 3 5 1 Methodology For Handling Waters cccsssssscccececeeeeeeeeeeeeeeeeaaanees 9 3 5 2 Specifying WOLCMS wicsnsececececccecedasecesec eves stun ddeesseeetenstundeesteeeteactesdseesees 10 3 6 Defining the Binding Site scx c ccc cece esesescsezcveneuve cess cach tacuessecess cach deeuevsedessenetene 12 3 6 1 O72 a i ee 12 3 6 2 Defining a Binding Site from an AtOM ccccccccccccesssssseeeeeeeeesseaaeees 13 3 6 3 Defining a Binding Site from a Point cccccccecceccecseesseeeseeeeesaeaeeess 14 3 6 4 Defining a Binding Site from a Reference Ligand ccccccceseeeeeees 16 3 6 5 Defining a Binding Site from a List of Atoms or Residues 068 17 3 6 6 Cavity D tectloniicecascteciscctee hi cateotenruresitecernetere ete reteretee tenets 18 3 6 7 Salvent Acce ssibility 2 202 Gn8e Sa Se eee oe Re Oe OO 19 3 7 Rotatable O H and NH3 Group ccccccccccesssseeececceseeaeesseseeeeeesaeaaaseeeeeeeeeeaaa 20 3 8 Docking into a Rigid Protein teroron niiina aiii oi 20 3 9 Metal OMS ee coc cecacccsccvscncberenovesvsvesucboveresesesevesvanshsedesesusodusapssedeeeduansusannescepevesd 20 3 9 1 Preparing a Protein Input
33. Accuracy and Speed with Genetic Algorithm Parameter Settings 23 2 Selection Pressure Each of the genetic operations crossover migration mutation see Operator Weights Migrate Mutate Crossover takes information from parent chromosomes and assembles this information in child chromosomes The child chromosomes then replace the worst members of the population The selection of parent chromosomes is biased towards those of high fitness i e a fit chromosome is more likely to be a parent than an unfit one The selection pressure is defined as the ratio between the probability that the most fit member of the population is selected as a parent to the probability that an average member is selected as a parent Too high a selection pressure will result in the population converging too early For the GOLD docking algorithm a selection pressure of 1 1 seems appropriate although 1 125 may be better for library screening where the aim is faster convergence Changes to genetic algorithm parameters should be made with care see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 23 3 Number of Operations The genetic algorithm starts off with a random population each value in every chromosome is set to a random number Genetic operations crossover migration mutation see Operator Weights Migrate Mutate Crossover are then applied iteratively to the population The parameter Number of Operations or maxops is
34. Add Definition as 4 Selection Help R Run GOLD Run GOLD In The Background Finish Cancel e When specifying a list of atoms the atom numbers as they appear in the input protein must be provided Multiple atom numbers are permitted on each line in the file It is therefore possible to re use an existing active site definition by using the list of active atoms printed in the protein log file Example file format is shown below GOLD User Guide 17 618 646 651 1344 1346 1995 1996 1997 1556 647 625 1570 957 599 1569 1499 493 1558 1559 156 1311 1328 1330 1978 1998 678 1549 1550 1553 497 1563 1562 1830 1327 1976 1977 1446 871 1603 677 955 959 958 1544 1542 1546 1545 1543 483 585 2054 1448 2056 256 2096 583 1058 895 912 1800 1798 887 1829 255 1158 1003 886 1799 949 1823 1824 1825 1826 1831 950 487 556 1803 1804 57 558 206 971 1842 956 970 995 996 205 1039 74 969 1839 1840 964 1832 267 208 982 1841 269 1856 983 1156 1868 1168 984 994 266 1167 1169 1154 1038 1040 203 75 1037 1036 207 370 1299 268 981 1853 967 1846 1837 368 369 360 1241 265 329 1239 1240 332 366 367 1297 1298 1243 When specifying a list of residues the residues can be extracted from any text file including a standard GOLD solution file GOLD writes the active site residues list to the solution files if output of rotatable hydrogens is turned on The following formatting restrictions apply The list must begin with the following tag on its own line
35. B Shoichet J Alvarez Taylor amp Francis CRC Press Boca Raton Florida USA 2005 Modeling Water Molecules in Protein Ligand Docking Using GOLD Marcel L Verdonk Gianni Chessari Jason C Cole Michael J Hartshorn Christopher W Murray J Willem M Nissink Richard D Taylor and Robin Taylor J Med Chem 48 6504 6515 2005 Using Buriedness to Improve Discrimination Between Actives and Inactives in Docking N M O Boyle S C Brewerton and R Taylor J Chem Inf Model 48 1269 1278 2008 DOI 10 1021 ci8000452 The use of protein ligand interaction fingerproints in docking S C Brewerton Curr Opin Drug Discov Devel 11 3 356 64 2008 Empirical Scoring Functions for Advanced Protein Ligand Docking with PLANTS O Korb T St tzle and T E Exner Journal of Chemical Information and Modeling 49 1 84 96 2009 http pubs acs org doi abs 10 1021 ci8002982z References dealing with GOLD validation 140 A New Test Set for Validating Predictions of Protein Ligand Interactions J W M Nissink C Murray M Hartshorn M L Verdonk J C Cole and R Taylor Proteins 49 4 457 471 2002 Improved Protein Ligand Docking using GOLD M L Verdonk J C Cole M J Hartshorn C W Murray R D Taylor Proteins 52 609 623 2003 Diverse High Quality Test Set for the Validation of Protein Ligand Docking Performance M J Hartshorn M L Verdonk G Chessari S C Brewerton W T M Mooij P N
36. Binding Site Protein 1acj hydrolase carboxylic esterase HOH150 3 toggle I spin Select Ligands Configure Waters Protein 1acj hydrolase carboxylic esterase _HOH151 3 toggle ifspin Ligand Flexibility Fitness amp Search Options Protein 1acj hydrolase carboxylic esterase _HOH152 3 GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing Help R Run GOLD Run GOLD In The Background Finish Cancel Running GOLD Dockings All waters toggled Click on Fitness amp Search Options You can see the gold conf file for this docking job uses the ChemScore scoring function Ensure the Allow early termination flag is set off i e that there is no tick in the tickbox and that the Generate diverse solutions flag is set on The Diverse Solutions Options button allows the diverse solutions settings to be configured Keep the settings at default values Now go to GA Settings the Genetic Algorithm parameters used are preset and are the slowest thus most accurate settings 100 000 operations Because allowing the waters to toggle on or off normally increases the size of the search necessary to find a good docking mode it is generally recommended to increase the search time allowed per ligand when toggling waters The search problem becomes harder the more waters that are included In this case because of the small size of the binding site and the fact the ligand has no rotatable bonds the se
37. CSD can be utilised by GOLD These distributions can be used to restrict the ligand conformational space sampled by the genetic algorithm e Using torsion angle distributions in this way will not make GOLD go any faster However it may improve the chances of GOLD finding the correct answer by biasing the search towards ligand torsion angle values that are commonly observed in crystal structures It may also improve convergence and so make improve accuracy with faster settings see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings e To enable the use of torsion angle distributions click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Use Torsion Angle Distributions check box e Atorsion angle distribution file must be specified Either enter the path and filename of the file or click on the button and use the file selection window to choose the file Two torsion angle distribution files are provided with GOLD gold tordist this is the default file GOLD User Guide 81 8 7 2 8 7 3 82 mimumba tordist this contains all the torsional distributions used in the MIMUMBA program Klebe and Mietzner J Comput Aided Mol Des 8 583 606 1994 It is possible to customise torsion angle distribution information by editing one of the standard torsion angle distribution files see Editing Torsion Angle Distribution Files Editing
38. Calculate and write out additional data after each docking Add extra terms to the scoring function Implement a completely new scoring function e Full documentation for the GOLD Scoring Function Application Programming Interface API is provided with the GOLD distribution UNIX SGOLD_DIR gold api_doc index html Windows lt InstallDir gt GOLD gold api_doc index html where lt InstallDir gt is usually C Program Files CCDC GOLD Suite e See GOLD Scoring Function Application Programming Interface API documentation GOLD User Guide 73 7 7 74 A good knowledge of the C programming language is required together with some experience in using GOLD Click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select User Defined Score from the drop down menu The Scoring Function Shared Object UNIX or Scoring Function DLL Windows is used to specify a path to a dynamically loadable shared object library Either enter the path and filename or click on the button and use the file selection window to choose the file GOLD uses shared objects or dynamically loadable libraries to allow new or modified scoring functions to be plugged in Two shared object files are relevant The main GOLD shared object which is called Libgold so UNIX or gold dll Windows The scoring function shared objects which by default are called libfitfunc_dll so UNIX gold
39. Coordinate and enter the orthogonal x y z coordinates of a single point upon which to position the sphere e Click on Done in the Edit Sphere dialogue once the sphere has been defined e A score contribution must also be specified This is the value that will be added to the fitness score for each specified non hydrogen ligand atom found within the GOLD User Guide 99 10 5 10 5 1 10 5 2 100 sphere region The total contribution added will therefore depend on the number of atoms located within the sphere Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor It is possible to define multiple region hydrophobic constraints Similarity Constraints This constraint can be used to bias the conformation of docked ligands towards a given solution or template Method Used for Similarity Constraints This constraint will bias the conformation of docked ligands towards a given solution This solution or template can for example be another ligand in a known conformation a common core useful when docking ligands of a combinatorial set or it may just be a large substructure that is expected or known to bind in a certain way The template must be supplied as a MOL2 file Unlike the distance based constraints which reduce the score for ligands that adopt unfavourable orientations this constraint will add an energy term to the score based on the similarity betwe
40. Diverse Solution Options P Use the internal ligand energy offset J Read hydrophobic fitting points File fie _pts mol2 yy view GOLD parameter file DEFAULT NN Edit Help R Run GOLD Run GOLD In The Background Finish Cancel The rms deviation takes account of any ligand symmetry Early termination does not always save as much time as you might think because it tends to be invoked for easy i e relatively rigid ligands which are quick to dock anyway Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings Genetic Algorithm Overview GOLD optimises the fitness score by using a genetic algorithm A population of potential solutions i e possible docked orientations of the ligand is set up at random Each member of the population is encoded as a chromosome which contains information about the mapping of ligand H bond atoms onto complementary protein H bond atoms mapping of hydrophobic points on the ligand onto protein hydrophobic points and the conformation around flexible ligand bonds and protein OH groups Each chromosome is assigned a fitness score based on its predicted binding affinity and the chromosomes within the population are ranked according to fitness The population of chromosomes is iteratively optimised At each step a point mutation may occur in a chromosome or two chromosomes may mate to give a GOLD User Guide child The selection of parent chromosomes is bia
41. M All Entries Z 1acm O Z Ligands Ma Wc Z Metals lt gt O chans IE Ligands Contact Management ax Define H Bonds Define Short Contacts Ligand Sphere Protein H Bonds Short Contac 1 1ACM J lt gt Graphics Object Explorer ax Explore non atomic graphics objects Right dick items for available options Entries ACM Certain groups can be represented in more than one way i e have more than one canonical form such as nitro carboxylate and amidinium In such cases there is usually a right and a wrong representation for use in GOLD The conventions used for some common difficult groups and further help on setting up the ligand is provided see Setting Up Ligands The phosphate and carboxylate groups are unprotonated and in each case the bond to the O atom has been assigned as aromatic In this case these groups have been correctly handled by Hermes and so no editing is required Had modifications been required editing functionality is available under Edit Edit Structure Re display all the Chains Ligands and Metals tickboxes so that the 3D view displays the entire protein structure In the Protein setup window click on the Delete Ligands option beneath the Extract Delete Waters option This window enables us to extract and delete a ligand from the protein in order to set it up for docking As we are going to be docking into chain A we need to first remove the co crystallised l
42. Mortenson C W Murray J Med Chem 50 726 741 2007 Pose prediction and virtual screening performance of GOLD scoring functions in a standardised test J W Liebeschuetz J C Cole O Korb J Comput Aided Mol Des 50 737 48 2012 DOI 10 1007 s10822 012 9551 4 GOLD User Guide General GOLD docking virtual screening papers that may be of interest GOLD User Guide Life science Applications of the Cambridge Structural Database R Taylor Acta Cryst D58 879 888 2002 Virtual Screening Using Protein Ligand Docking Avoiding Artificial Enrichment Marcel L Verdonk Valerio Berdini Michael J Hartshorn Wijnand T M Mooij Christopher W Murray Richard D Taylor and Paul Watson J Chem Inf Comput Sci 44 793 806 2004 Comparing protein ligand docking programs is difficult Jason C Cole Christopher W Murray J Willem M Nissink Richard D Taylor Robin Taylor Proteins 60 325 332 2005 Evaluating docking programs keeping the playing field level J W Liebeschuetz JCAMD Vol 22 No 3 4 2008 229 238 Testing Assumptions and Hypotheses for Rescoring Success in Protein Ligand Docking N M O Boyle J W Liebeschuetz J C Cole J Chem Inf Model 49 1871 1878 2009 DOI 10 1021 ci900164f Docking performance of fragments and drug like compounds M Verdonk I Giangreco R Hall O Korb P Mortensen C Murray J Med Chem 54 5422 5431 2011 DOI 10 1021 jm200558u 141 19 Acknowled
43. N_BINS is the number of bins used in the torsion histogram REMOVE_HIGH_ENERGY and DELTA_E are parameters that can be used to control the filtering out of high energy torsion angles If torsion angle distributions are used GOLD will no longer sample over 360 degrees but will constrain the torsion to values contained in the histogram However if a histogram contains a large number of entries there may be some high energy torsions within the histogram GOLD therefore provides a method for filtering out such high energy torsions set REMOVE_HIGH_ENERGY 1 and DELTA_E Eto remove those bars in the histogram that correspond to torsions that are E kcal mol higher in energy than the most populated state The ground state of the torsion is assumed to correspond to the maximum peak in the torsional histogram The energy difference between this ground state and any other peak in the torsion angle histogram is then assumed to be approximately given by the partition function The following table indicates the relationship between the value of DELTA_E and the ratio high low where high is the height of the biggest bar in the histogram and low is the height below which bars will be removed from the histogram DELTA_E Ratio 3 0 161 2 5 69 2 0 30 For example if REMOVE_HIGH_ENERGY 1 and DELTA_E 2 5 those bars which are 1 69th or less of the height of the largest bar will be removed from the histogram and torsion angles corresponding to these bars will
44. Program Settings in Configuration Files The configuration file is a text file which specifies the GOLD calculation that is to be run including details of the ligand the protein binding site the fitness function parameter file to be used the torsion distribution file to be used and the genetic algorithm parameters Although the file can be generated with a standard text editor the easiest way to create it is to use the GOLD front end Any settings that have been defined in the GOLD interface can be saved as a configuration file by selecting the Save button located next to the Conf file entry box at the top of the GOLD Setup window Alternatively you will be prompted to save the file if you start a GOLD job from the interface by selecting either Run GOLD or Run GOLD in the background By default the configuration file will be saved in the directory from which GOLD was opened and will be called gold conf Use the Conf file entry box at the top of the GOLD Setup window to change the file name and or directory any file name can be used Once a configuration file has been created it can be re used either as a quick way of reading program settings into the GOLD front end or to run GOLD from the command line see Running GOLD To load a previously created configuration file into GOLD interface enter the file name into the Conf file entry box at the top of the GOLD Setup window Alternatively click on the Load button and use the file sel
45. Protonation and Tautomeric States Active waters i e those that you would like GOLD to consider during docking must be provided in separate files one water molecule per file Read in each of the water molecule files HOH150 mol2 HOH151 mol2 and HOH152 mol2 Read in the file ligand mol2 from the folder containing the tutorial15 files You will be able to see how the ligand tacrine chooses to bind Measuring distances in Hermes you will find that the waters HOH150 and HOH151 are within hydrogen bonding distance of the NH of the ligand as well within hydrogen bonding distance of hydrogen bond acceptors on the protein The third water is at a position where it can make a hydrogen bond to the same histidine backbone carbonyl as the protonated ring nitrogen of the ligand This water cannot be accommodated if tacrine takes up its normal binding mode None of the hydrogen positions on the waters have been positioned for optimal hydrogen bonding This doesn t matter as the water hydrogen positions can be optimised during docking 179 Close all the files by clicking on File then Close All Files Setting up protein bound waters 180 A configuration file gold conf has been provided for this tutorial which will automatically load most of the settings and parameter values for this tutorial into the GOLD front end From within Hermes click on GOLD then Setup and Run a Docking in the top level menu Load the gold conf for tutorial 5 by s
46. The RMSDs for the top ranked pose in each cluster compared to the native ligand pose in the example above are cluster 1 7 35 A GoldScore fitness 29 48 ranked 1st cluster 2 5 20 A GoldScore fitness 26 19 ranked 3rd cluster 3 0 82 A GoldScore fitness 25 23 ranked 7th cluster 4 7 82 A GoldScore fitness 24 48 ranked 9th e You will observe something similar in the docking you have carried out Conclusions e The binding site of 3MTH is large In addition there are a relatively small number of donor and or acceptor points in the active site where a ligand might bind Furthermore the co crystallised ligand methylparaben insulin also contains few functional groups All of these factors mean the docking of the native ligand back into 3MTH is a complex problem for GOLD e Enabling the diverse solutions feature produces a number of different docked poses two of which are found to be close to the native ligand pose This ends the tutorial 202 GOLD User Guide 20 9 Tutorial 9 Running a Covalent Docking Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial9 toa directory to which you have write permissions The object of this tutorial is to perform a covalent docking using 1ASE taken from the CCDC Astex validation test set This is available to download as GOLD Validation Sets from http www ccdc cam ac uk SupportandResources Downloads pages ProtectedDow nloadProd
47. Torsion Terms e ASP _GRID_ SPACING Default 0 3 Parameter to control the density of the pre calculated grid used for evaluation of the atom atom potentials see The Generation of Potentials e ASP GRID INTERPOLATE Uncomment the ASP_GRID_INTERPOLATE to more accurately calculate the distance between atoms for the calculation of the score Grid points surrounding atoms are used to interpolate a more exact atom location This will give a similar effect as increasing the grid density e ASP MAX DISTANCE Default 6 0 The max distance set for interaction between ligand and protein atoms e ASP _GRID_ LOOKUP This setting is on by default the score is evaluated from the generated grid points e ASP DIRECTORY Default DEFAULT Sets the location of the ASP potentials by default the location is GOLD_DIR gold asp tables e TARGETED ASP DIRECTORY Uncomment this parameter to specify the location of customised targeted ASP potentials see Targeted Scoring Functions e SAVE ASP MAPS Default 0 If set to 1 the map generated for each ASP ligand atom type is printed out 7 6 User Defined Scoring Function e In addition to the choice of scoring functions provided i e GoldScore ChemScore ASP and CHEMPLP users can also implement their own scoring function e The GOLD scoring function Application Programming Interface API allows users to modify the GOLD scoring function mechanism in order to
48. VdW clash component of the GoldScore for one or more residues in the protein We will examine the docking of a ligand to two different crystal structures of Estrogen Receptor Alpha The structures differ in that a small loop movement constrains the binding site of one of the structures pdb code 1x7r slightly more than for the other structure pdb code 112i The figure below shows the superposition of both protein structures 1x7r corresponds to the protein colour coded light blue and the ligand colour coded green 112i corresponds to the protein colour coded orange with the ligand colour coded yellow 194 Most of the binding site is well superimposed however above the ligands you can see that there is movement of a protein loop that brings Leu346 closer in to the ligand in 1x7r than in 112i This superposition suggests that a clash would exist if the GOLD User Guide ligand from 112i were docked into 1x7r This might prevent the correct binding mode being rated highly if using a scoring function such as GoldScore with a clash term that increases sharply with proximity to the protein Other residues such as Met343 also do not superimpose well as a consequence of this loop movement However these residue shifts appear to have less of an impact on the size of the active site than does that of Leu346 You can view this superposition yourself by opening Hermes or another protein visualiser and reading in the file 1x7r_1 2i_sup mol2 Docki
49. a Gaussian smoothed block function see Block Functions in ChemScore whose purpose is to reduce the contribution of the metal acceptor interaction if the geometry is not ideal z 2 B TM Ris R O ag All ligand All protein acceptors metals e The table below describes the various parameters in this equation their meanings and what they are called in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File Metal binding parameters in ChemScore Term Meaning Name in ChemScore File Default Value TaM The actual acceptor metal distance in Calculated for each acceptor metal pair Rideal The ideal acceptor metal distance METAL R1 23 6 Rmax The maximum acceptor metal distance to be METAL R2 3 0 considered a binding interaction O metal The Gaussian smearing sigma associated with METAL R_SIGMA 0 1 this term e The metal binding term has a regression coefficient associated with it v2 By default this is set to 6 03 The name of this coefficient in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File is METAL COEFFICIENT e The lipophilic term is defined in a similar way Bis a 2 2 B ly Riga Ks O lipo Allligand All protein lipophilic atoms lipophilic atoms e The table below describes the various parameters in this equation their meanings and what they are called in the Che
50. a particular side chain at it s input conformation i e to make it non flexible during docking click on the Rigid button Any previously defined rotamers will be lost Setting a side chain to be freely rotatable To allow a side chain to rotate freely during docking click on the Free button This will define a single rotamer where all rotatable torsions are permitted to vary over the range 180 to 180 Any previously defined rotamers will be lost GOLD User Guide From a rotamer library The file lt GOLD DIR gt gold rotamer_library txt contains information taken from the paper The Penultimate Rotamer Library S C Lovell J M Word J S Richardson amp D C Richardson Proteins 40 389 408 2000 It is a compilation of the most commonly observed side chain conformations for the naturally occurring amino acids To define rotamers corresponding to these commonly observed side chain conformations click on the Library button Note that the library settings are simply a starting point users are encouraged to generate their own rotamers for optimal results From the protein input file Click on the Crystal button to define a rotamer in which all rotatable torsions in the side chain will be allowed to vary over the range delta chi to delta chi where chi values are taken from the protein input file 4 1 4 Deleting and Editing Rotamer Definitions GOLD User Guide To remove or copy a particular rotamer definition right
51. assign if you are setting atom types manually Functional Group 2D Diagram Notes Amidinium H N pl3 H m N pI3 H H Carboxylate O co2 ary C 2 ars O co2 Enolate phenolate O co2 oxygen C 2 ue WN id apasal ar Guanidinium H C N pI3 H ars N pl3z C cat ar sar H N pI3 H H GOLD User Guide 51 Functional Group 2D Diagram N oxide 0 2 Nitro 0 2 C N pl3 0 2 Nitrogen anionic N pl3 Nitrogen cationic aromatic L 8 e N al SS ar A C Notes For example an anionic imidazole ring would be 62 R N pl3 So27 C 2 N 2 For example the pteridine ring system in methotrexate PDB code 4DFR would be NH2 N K J N N pI3 NH H GOLD User Guide Functional Group 2D Diagram Notes 0 3 A serine protease transition state analogue example is shown Oxygen anionic protein C 0 co2 X Phosphate bridging 0 3 0 3 FOR ee ON C 2 C ars ar gt O co2 O co2 Phosphate terminal 0 3 ar O co2 ANL C P 3 ars ar AX O co2 O co2 Sulphonamide GOLD will treat the nitrogen atom as 02 02 a planar trigonal nitrogen i e not W capable of accepting a hydrogen S02 c bond However pyramidal VART ERR sulphonamide nitrogen atoms are C N pl3 now typed as N 3 if the geometry read into GOLD is pyramidal rather H than N pl3 and are treated as H bond acceptors i e they have a fitting point allowing them to coordinate metal groups Sulphonate O co2 ap C
52. atom 2548 Soft Potentials Metals SEA Ligand link mode Atom Substructure Interaction Motif Substructure link atom 2 Substructure file Ints and Settings henderson Desktop substructure mol2 s IV Use topology matching to check test equivalent atoms You can define the covalent link atoms by right clicking in the viewer or by typing in the edit box The ligand link atom can be defined either by a single atom in the ligand or by an atom in a substructure that can be matched against multiple ligands IF using a substructure you must enter the substructure file Help R Run GOLD Run GOLD In The Background Finish Cancel To specify the Substructure file either enter the path and filename of the file or click on the button and use the file selection window to choose the file Define both the Protein link atom and Substructure link atom This can be done by clicking on an atom in the visualiser Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box Enable the Use topology matching to check equivalent atoms check box if the constraint refers to a substructure atom and therefore a ligand atom which is topologically equivalent to other atoms e g it is one of the oxygen atoms of an ionised carboxylate group GOLD will then use whichever of the equivalent atoms gives the best result 5 8 Specifying a L
53. atom in question will be replaced with a dummy atom type Du If this is the case a warning message will be given in the gold_protein log file The presence of dummy atoms should not significantly affect the docking prediction since dummy atoms are neither considered as donors or acceptors Dummy atoms may be visualised in Hermes by activating the Show unknown atoms tickbox on the hermes top level menu Atom types may be set manually provided you are using MOL2 input files see Manually Setting Atom and Bond Types Alternatively they can be set automatically see Automatically Setting Atom and Bond Types Unless you are an expert GOLD user or are dealing with a very unusual ligand structure you are recommended to use this option However you still need to input the ligand and protein structures correctly e g with correct bond orders and appropriate protonation states 6 2 Automatically Setting Atom and Bond Types GOLD User Guide Unless you are an expert GOLD user or are dealing with a very unusual ligand structure you are recommended to use the automatic atom type assigner To automatically set atom and bond types click on Atom Typing from the list of Global Options given on the left of the GOLD Setup window Atom and bond types can then be assigned automatically for the ligand and or protein by switching on the appropriate check box es GOLD assigns atom types from the information about element types and bond orders in the in
54. available through which the link can be made If the link atom on the ligand does not have a free valence having a hydrogen instead then the docking will proceed and the hydrogen will be ignored in terms of its contribution to the fitness score It will however still be displayed when docking poses are visualised Inside the GOLD least squares fitting routine the link atom in the ligand will be forced to fit onto the link atom in the protein In order to make sure that the geometry of the bound ligand is correct the angle bending potential from the Tripos Force Field has been incorporated into the fitness function On evaluating the score for the docked ligand the angle bending energy for the link atom is included in the calculation of the fitness score The Tripos force field is described in Validation of the General Purpose Tripos 5 2 Force Field M Clark R D Cramer IIl amp Nicole Van Opdenbosch J Comp Chem 10 982 1989 This seems to work well in the systems on which GOLD was validated However since the protein is held rigid apart from hydroxyl hydrogen atoms it does require that the position of the link atom in the protein is sensible 5 7 2 Setting Up a Single Covalent Link GOLD User Guide Set up the protein and ligand structures so that they both contain the link atom see Method Used for Docking Covalently Bound Ligands Covalent constraints are specific to the protein thus click on the protein tab e g Prote
55. bond by a ligand hydrophobic group other close contacts between protein and ligand the buried surface area of the ligand or of certain types of atoms in the ligand e g hydrophobic atoms whether particular regions of the binding site are occupied by the ligand simple properties such as the number of H bonding ligand atoms molecular weight of ligand number of rotatable bonds For further information on Visualising and Refining Selections of Docking Poses and on Defining and Calculating Descriptors refer to the Hermes user guide Analysing Results in GoldMine Overview of GoldMine GoldMine is a tool for the analysis and post processing of docking results It is possible to create within GoldMine a database of docking data which may comprise one or more sets of docking data A GoldMine Database or GoldMine DB is the term we will use to describe such a database GoldMine can be used to combine and analyse several docking runs For instance docking runs against different protein models may be combined within a GoldMine GOLD User Guide DB and analysed for selectivity and specificity Docking runs carried out against one protein model but scored using different scoring functions may also be combined within a GoldMine DB Each set of docking results saved within a GoldMine DB will contain one or multiple binding poses for each ligand and the corresponding protein configurations They will also contain any nume
56. bonds list written out at the end of the docking run when the chromosome is decoded e g Chromosome decoded Ligand Torsions 61 41 40 3 51 44 40 41 61 63 165 82 matched torsion ester 26 57 48 38 60 63 65 64 63 61 71 33 matched torsion acid T1 67 65 64 63 161 95 matched torsion acid T2 In some cases a rotatable bond may match more than one torsion angle distribution If this happens a score is calculated for each torsion angle distribution and the distribution with the highest score is selected Note A weighting scheme is used when matching rotatable bonds in the ligand to a torsion angle distribution such that more specific torsion definitions are taken in preference to more generic ones Each portion of the torsion angle distribution contributes to the score as follows GOLD User Guide Element atom type 1 5 SYBYL atom type 2 0 Fragment 3 0 Hydrogen count 2 0 Bond linkage 0 5 8 8 Overriding Automatic Bond Settings GOLD User Guide When using ligand flexibility options e g Flip amide bonds see Flipping Amide Bonds or Flip all planar R NR1R2 see Flipping Planar Nitrogens the bond in question is treated in a specific manner at ligand initialisation to prepare it for the docking run in both the aforementioned cases the bond is flattened at ligand initialisation prior to it being flipped during docking If a bond is e g desired to rotate freely rather than flip during docking this fine grain
57. check box 15 1 10 File Containing Error Messages e The file gold err lists any errors found by the program These are generally fatal and cause the program to stop It is a good idea to check gold err if something goes wrong e Errors and warnings generated by the atom type checker are also written to gold err If you are unsure about your atom typing you should therefore check this file For example Ligand in file ligand mol2 named LIM AL5_ 555 pdbibni_ i starting at address 190 raised the following warnings and or errors Warning message check_atom types atom 21 in ligand mol2 is type N am resetting to type N pls e In the parallel version warning messages are logged in individual error files one for each process They are not sent back to the central parallel scheduling process e gold err is line buffered so errors are logged immediately 15 1 11 Process File e The file gold pid records the user host and process number of the GOLD job It is deleted when GOLD exits Its purpose is to stop the user running two GOLD jobs in the same directory 130 GOLD User Guide If the machine goes down or GOLD crashes or is killed with signal 9 you will need to remove gold pid before you can run another GOLD job in the same directory 15 1 12 Seed Log File A file called gold seed_log is written to the output directory for each docking run GOLD uses a random number generator for some operations e g choosing which genet
58. click on the rotamer name in the Edit Rotamer Library dialogue and select either Delete rotamer or Copy rotamer from the resulting drop down menu To edit a particular rotamer definition right click on the rotamer name in the Edit Rotamer Library dialogue and select Edit this rotamer from the resulting drop down menu This will open the Edit rotamer dialogue Name Rotamert sts chit 49 fio ce 10 fo Energy fo Rotamer angles should be in the range 180 to 180 Rotamer deltas should be a single number in the range 0 to 180 or a pair of the form 10 20 to give an asymmetric range Energy is the energy penalty to be applied when this rotamer is selected cel The rotamer Name Chi and corresponding Delta values can be changed by typing into the appropriate entry boxes Chi values should be a single number in the range 180 to 180 Delta values should be a single number in the range O to 180 or a pair of numbers of the form x y to specify an asymmetic range From dials Rotamers can be specified directly To set a chi value click on the dial and while holding down the mouse button move the red indicator line to the required position The corresponding torsion will rotate within the Hermes visualiser to show the current value Alternatively type the required chi value into the entry box directly under the dial Once the chi and delta values have been set click on the From Dials button to add this rotamer definition 2
59. distance range Click on Substructure within the Constraints tree to open the substructure constraint set up window We now need to select the atoms involved in the constraint specifically the zinc atom and the N atom in the substructure moi2 file The zinc atom is coloured grey in the 3D view Click on the metal atom the metal atom will be highlighted with a cyan sphere and the atom ID 2041 will be entered into the Protein atom number window A substructure file substructure mol2 containing a sulphonamide group has been provided for this tutorial and can be found in the folder to which you copied the tutorial4 files When creating your own substructure files it is recommended that you set atom types manually see Setting Up Substructure Based Distance Constraints since an incomplete fragment can cause problems with automatic atom typing Click on the Substructure file button then select the file substructure mol2 from the folder to which you copied the tutorial4 files and hit Open This automatically loads the substructure file into the 3D view If the substructure is not clear you can suppress the protein atoms via the tickbox adjacent to Protein 1cil lyase oxo acid in the Molecule Explorer Select the N atom in the substructure This will enter the atom ID 4 into the Substructure atom no dialogue The N atom will also be surrounded by a cyan sphere Specify the allowed range of separation by entering a Minimum separation of 1
60. e a point see Defining a Binding Site from a Point or a reference ligand see Defining a Binding Site from a Reference Ligand It is not possible to define the active site using an atom or a list of atoms or residues e Ligands are specified for ensemble docking in the same way as when docking into an individual protein see Specifying the Ligand File s A maximum of 20 proteins can be specified in an ensemble e The following setup options can be applied across the entire ensemble Specification of active waters see Specifying Waters Waters should not be considered to be associated with a specific protein rather representative waters should be specified that will be used in all proteins GOLD User Guide 35 4 3 4 36 Similarity constraint see Setting Up a Similarity Constraint Scaffold constraint see Setting Up Scaffold Match Constraints Region constraint see Setting Up Region Hydrophobic Constraints Settings specific to each protein are controlled from within the individual protein tabs i e Protein setup addition of H atoms extraction deletion of water molecules extraction of ligands Specification of flexible side chains see Side Chain Flexibility Note that the active site definition must be set before flexible side chains can be setup Special treatment of metal atoms see Metal lons Distance constraint see Setting Up a Distance Constraint Substructure constraint see Set
61. facilitate protein superimposition Proteins can be overlaid by matching residues based on label matching residues based on sequence number or by matching residues based on sequence alignment Optionally a component of fasta called ggsearch2 can be used for sequence alignment of proteins to be superimposed The package can be downloaded from http fasta bioch virginia edu fasta_www2 fasta_down shtml In both cases above the wizard guides you through the superimposition process 4 3 3 Setting up an Ensemble Docking 34 All proteins currently loaded into Hermes are listed in the Proteins window and can be selected or deselected for use in the ensemble using their associated tick box Only proteins are listed in this dialogue To view all other loaded files activate the List all loaded files not just proteins tick box Each loaded protein has its own tab adjacent to the Global Options tab labelled with the name taken from the protein file e g pdb1qpc_full_entry below GOLD User Guide gt GOLD Setup BBE Conf file C Documents and Settings henderson Desktop ensemble files ensemble conf Load Save Global Options pdbigpc_full_entry pdbigpd_full_entry pdbigpe_full_entry pdb1igpj_full_entry pdb3ick_a l wizard Templates Select proteins to use Load Protein Superimpose Proteins Proteins Define Binding Site pdbigpc_full_entry Select Ligands Configure Waters pdb1gpd_full_entry Ligand Flexibility p
62. files are written Click on the Run GOLD button In the Finish GOLD Configuration window you will be prompted that the GOLD configuration has been updated and needs to be saved Change the configuration file name to e g gold_covalent conf then hit Save to start the GOLD run Analysis of output GOLD User Guide Once the docking has completed click on the gold_ligand_m1 log text in the Run GOLD window This will load the ligand log file into the Run GOLD window enabling us to view the progress of each genetic algorithm run Scroll through this file You will notice that as the ligand was being initialised the covalent constraint was analysed 205 206 gt Run GOLD 21 xi list of ligand logs gold log gold_protein log gold err Messages gold_ligand_m1 log Protein atom id Ligand atom id Angle term entry 178 H N acid C 2 120 0 Angle term entry 178 C 3 N acid C 2 120 0 N ple donors pro L Acidic nitrogen acceptors none found Sulfur acceptors none found 0 3 atom 12 is not an acceptor skipped 14 POCOZ acceptor 15 POCOZ acceptor 16 POCOZ acceptor Donor atoms 6 Cen er AN This log file will be updated every 2 seconds Interrupt G View Solutions Close You will notice a couple of Angle terms these correspond to the geometry around the link N atom During docking GOLD refers to angle and torsion data in its parameter file to ensure the pose s adopted by the ligand are c
63. flexible conf Load Save Options Protein 1fax coagulation factor Waters Delete Ligands Soft Potentials Metals Constraints Covalent Help R Protonation amp Tautomers Flexible Sidechains Interaction Motif Rotamer library Program Files CCDC GOLD Suite GOLD gold rotamer_library txt sei TYR99A Rigid 45P100 4 Rigid ASP102 A Rigid ARG143 4 Rigid GLU147 A Rigid SER172 4 Rigid SER173 4 Rigid PHE174 A Rigid ILE175 4 Rigid ILE1764 Rigid MET180 4 Rigid CY5182 A Rigid ASP189 A Rigid CY5191 A Rigid GLN192 4 Constrained 9 rotamers ASP194 A Rigid SER195 A Rigid ILE212 A Rigid VAL213A Rigid xl Highlight ai Highlight None Highlight Flexible Edit Choose which residues of the binding site should be flexible and set rotational parameters For them Run GOLD Run GOLD In The Background Finish Cancel e By default all side chains will be treated as rigid i e they will be held fixed at their input conformation during the docking To make a side chain flexible you can either Select the side chain by clicking on it in the list Once selected a side chain will be highlighted in the Hermes visualiser Once a side chain has been selected you will be required the define one or more allowed rotamers Each rotamer specifies the torsion angles that are permitted to vary and the allowed values or ranges of values for those torsion angles Click o
64. is a tabbed view that allows you to inspect various files that are written while the docking proceeds Once the job is complete the message Finished Docking Ligand ligand mol2 will appear in the gold_ligand_m1 log tabbed view Once the GOLD job is complete load the results into Hermes using the View Solutions button 169 Analysis of Output 170 Inspect the go d_protein log file by hitting the gold_protein log tab in the Run GOLD window If you have already closed the Run GOLD window this file can be found in the output directory specified see The GOLD Configuration File and can be read using a text editor This file contains details of the protein initialisation Now return to the list of ligand logs window and click on gold_ligand_m1 log again if you have closed the Run GOLD window this information can be found in the gold_ligand_m1 1og file stored in your specified output directory This file gives a total fitness score and a breakdown of the fitness into its constituent energy terms for each docking performed on the ligand A constraint scoring term DE con is listed for each docking If a solution predicted by GOLD satisfies all of the protein H bond constraints then the contribution from this scoring term will be 0 00 However for solutions in which not all of the constraints are satisfied a penalty will be applied to the fitness score for each constrained H bond that is not formed The value of this penalty is the Constraint
65. is also used for ChemScore this is called asp params Parameters within this file can also be modified see Altering ASP Fitness Function Parameters the asp params File Reference State The ASP scoring function differs from other statistical potentials by the choice of the so called reference state The reference state is the expected number of contacts if there were no interaction between the atoms i e at long distances incorporating any corrections The reference state determines how the raw distribution of observations is transformed into potentials 69 7 5 3 70 ASP _ obs nexp Contacts between atoms are usually determined by radial distribution functions RDFs Given an atom at some position the RDF will tell us how many other atoms we can expect to find at a distance between r to r dr where dr is the bin width in the RDF and can be thought of as the thickness of a spherical shell A statistical potential between two atom types i and j is defined as n i j r _ obs gt StatScore i j r In _statScore n 1 r exp Where the denominator is the reference state of the potential for ASP the reference state is given by ll ij r r 8 0 oa cn a f ir fG r Amr Ar fli PAG Anr Ar _ 6 9 The average contact density is taken to be the average between 6 0 and 8 0 of the corrected RDF At this long range atoms are not considered to make any specific interactions and should ensure that
66. it appears in the scaffold file directly into entry box Limiting the number of atoms to be matched can be useful for large rigid scaffolds In such a GOLD User Guide case specifying only a few atoms distributed throughout the scaffold can be sufficient to obtain a good 3D superimposition Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor 10 7 Interaction Motif Constraint This constraint can be used to bias the docking towards solutions that form particular protein ligand binding motifs This constraint could be used e g when there is experimental evidence such as a number of X ray structures from a fragment screen which show that certain combinations of interactions motifs are commonly formed by groups of fragment binders Such motifs can therefore be considered as favourable and this information can be incorporated into the docking in order to bias the ligand poses that are generated For further information see The use of protein ligand interaction fingerproints in docking see References 10 7 1 Method Used for the Interaction Motif Constraint GOLD User Guide Interaction motif constraints are applicable to individual protein ligand complexes i e must be set up individually for each protein ligand if performing ensemble docking This constraint is used to bias the docking towards solutions that form particular protein ligand binding motifs One
67. ligand a bond is considered frozen if one or more atoms on both sides of the rotatable bond is in contact with the protein The expression is deemed to have a value of zero if there are no rotatable bonds in the ligand P r and P r are the percentages of non hydrogen atoms on either side of the rotatable bond that are not lipophilic For example if there are 10 non hydrogen atoms on one side of the bond of which 3 are not lipophilic and there are 20 non hydrogen atoms on the other side of which 2 are not lipophilic then P r and P r are 30 and 10 respectively The regression coefficient associated with this term v4 has the default value 2 56 The name of this coefficient in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File is ROT COEFFICIENT GOLD User Guide 7 4 6 Clash Penalty and Internal Torsion Terms GOLD User Guide Clashes between protein and ligand atoms and ligand internal torsional strain are accommodated by penalty terms These terms are included to prevent poor geometries in docking The clash penalty terms in ChemScore differ on the nature of the contact i e whether it is a hydrogen bonding contact a metal binding contact or neither of these Any hydrogen bond with an H A distance shorter than Pppong A contributes a clash term of _ 20 0 x ee r clash hbond Se x Seal The value of rpbong default 1 6A
68. manipulates a pool of chromosomes of size popsize Number of Islands The size of this pool should be such that the optimisation converges within the specified maximum number of operations Number of Operations If the pool size is too small for a given value of Number of Operations the algorithm will converge prematurely Conversely if the pool size is too large the algorithm will terminate before it has converged e The annealing parameters van der Waals and Hydrogen Bonding allow poor hydrogen bonds to occur at the beginning of a genetic algorithm run in the expectation that they will evolve to better solutions Both the vdw and H bond annealing must be gradual and the population allowed plenty of time to adapt to changes in the fitness function e Because of these factors it is difficult to set GA parameters by hand and you are recommended to use automatic ligand dependent GA parameter settings see Using Automatic Ligand Dependent Genetic Algorithm Parameter Settings or one of the default parameter sets offered see Using Preset Genetic Algorithm Parameter Settings GOLD User Guide 109 11 3 3 110 Using Automatic Ligand Dependent Genetic Algorithm Parameter Settings The number of genetic operations performed crossover migration mutation is the key parameter in determining how long a GOLD run will take i e this parameter controls the coverage of the search space GOLD can automatically calculate an optimal number
69. never be sampled by the genetic algorithm The relationship between DELTA_E and ratio based on the partition function is ratio exp DELTA_E 0 5898 25 2 Format of Torsion Angle Distributions GOLD User Guide Each torsion angle distribution entry comprises three lines the first line is the name of the torsion angle the second line is the definition of the torsion angle the third line is the histogram The histogram should be a list of space separated integers The ith integer should be the number of observations in the torsion angle range of the ith bin There should be N_BINS integers in all The first bin starts at 180 degrees and the last bin ends at 180 233 234 e Torsion angle distributions are defined using Backus Naur Form BNF grammar as follows all the symbols in the table are part of the grammar except for which is used to indicate alternative fields TORSION DIRECTIVE NODE NEIGHBOURS NEIGHBOUR_NODE HYDROGENS ATOM FRAGMENT ATOM_DEF TYPE_DEF LINKAGE SYB_TYPE EL_TYPE NODE NODE NODE NODE NODE NODE NODE NODE DIRECTIVE NODE NODE NODE NODE DIRECTIVE DIRECTIVE expand lt min gt lt max gt period lt min gt lt max gt ATOM ATOM NEIGHBOURS NEIGHBOUR_NODE NEIGHBOUR_NODE NEIGHBOURS NODE HYDROGENS OH 1H 2H 3H ATOM_DEF ATOM_DEF FRAGMENT ribose adenine uracil benzene TYPE_DEF LINKAGE amp ltno space amp gtTYPE_DE
70. notice that the protein C atoms are coloured grey i e coloured by atom type while the C atoms of any ligands are coloured green Also in the Hermes 3D view you will notice the Molecule Explorer off to the left hand side of the Hermes interface Click on the adjacent to 1ACM and underneath All Entries The protein has been broken down into its constituent parts specifically Chains Ligands Metals and Waters Each of these has a corresponding adjacent to it Each successive time the is clicked on the component it corresponds to is broken down further In this way it is possible e g to identify specific protein residues or atoms in a ligand Display styles colours and labels and selection options are available by right clicking You will notice that the protein consists of 4 chains A B C and D The chain we will be focusing on is chain A In the GOLD Wizard click on the Next button to proceed to the Protein setup step Click on the 1ACM tab adjacent to the Global Options tab From under the JACM tab we can make some essential modifications to the protein specifically we can GOLD User Guide Add hydrogens see Adding Hydrogen Atoms all hydrogen atoms must be present in the protein input file see Protonation and Tautomeric States The hydrogen atoms are placed on the protein in order to ensure that ionisation and tautomeric states are defined unambiguously Advanced options within GOLD allow for switching b
71. of an ionised carboxylate group then GOLD will compute the constraint term using whichever of the equivalent atoms gives the best value automatically e Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor 10 2 2 Method Used for Substructure Based Distance Constraints e Substructure based constraints are applicable to individual protein ligand complexes i e must be set up individually for each protein ligand if performing ensemble docking e Itis possible to apply a distance constraint to multiple ligands which have a common functional group e The constraint forces GOLD to limit the distance between a protein atom and one atom of this functional group Docking solutions will be biased towards the specified distance range e During docking the constraint will be applied to any ligands which contain the specified substructure matching is performed on the basis of the element types and 2D connectivity and the resulting solutions will be biased towards the specified distance range GOLD always accounts for topology in the substructure e Note The substructure must be a sub graph rather than a complete molecule GOLD User Guide 93 10 2 3 94 As with normal distance constraints see Setting Up a Distance Constraint the score is reduced for unfavourable ligand solutions The amount of decrease in the score is determined by a weight term that the user must supply
72. of operations for a given ligand thereby making the most efficient use of search time e g small ligands containing only one or two rotatable bonds will generally require fewer genetic operations than larger highly flexible ligands The criteria used by GOLD to determine the optimal GA parameter settings for a given ligand include the number of rotatable bonds in the ligand ligand flexibility i e number of flexible ring corners flippable nitrogens etc see Ligand Flexibility the volume of the protein binding site and the number of water molecules considered during docking see Water Molecules The exact number of GA operations contributed e g for each rotatable bond in the ligand are defined in the gold params file see Altering GOLD Parameters the gold params File To enable automatic i e ligand dependent GA settings click on GA Settings from the list of Global Options given on the left of the GOLD Setup window then switch on the button labelled Automatic GOLD User Guide 3 GOLD Setup me E Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Wizard Templates Automatic Proteins Define Binding Site Preset Select Ligands User defined Waters Ligand Flexibility Fitness amp Search Options Output Options GoldMine Parallel GOLD Constraints Atom Typing E 1 1 1 1
73. of the Ca atom This is defined as a rotation of the improper torsion defined by the atom sequence CA N C CA To define an improper torsion enable the Improper check box in the Edit Rotamer Library dialogue An additional Improper torsion angle dial will become available for defining rotamers see Defining Rotamers In the example shown below an additional improper torsion has been specified The specification for the improper torsion angle will allow a rotation of or 30 degrees around the N C vector the zero angle corresponding to the Ca position given in the protein input file 31 4 1 6 32 Edit Rotamer Library TYR99 TYRSS Improper Chil Chi2 0 0 0 90 90 90 90 90 g0 180 180 180 0 J 106 i J51 Reset Reset Reset m Rotamer Library Operations Library Rigid Free Crystal From renee M Improper Delta Rotamert 0 Cancel It is not easy to decide on suitable rotation limits for improper torsions a trial and error approach is normally required but they often need to be quite large For example an improper rotation of about 40 degrees has to be applied to Tyr370 of 1dx4 for it to be possible to overlay the side chain closely onto the 1qon Tyr370 position Protein Protein Clashes By default when a flexible side chain is moved during docking GOLD checks whether any of its atoms clash with atoms in neighbouring residues This gives ri
74. possible to either Keep all solutions Keep the best n solutions for each ligand where n is a user specified number e g n 3 or Keep the top ranked solution for the best m ligands only i e retain just the best solution for only those m ligands with the best fitness scores where m is user specified e g m 100 e In addition you can filter out all solutions with fitness scores lower than a specified value by switching on the button labelled Reject solutions with fitness lower than x This filter can be used in combination with the options listed above For example you could save 3 solutions for each ligand and not keep any solution with a fitness lower than 50 GOLD User Guide 121 14 Running GOLD 14 1 Required Input Files The following files must be available before a GOLD job can be run One or more files containing the ligand s to be docked in MOL2 MOL SDor PDB format but PDB format is not recommended for ligand files see Setting Up Ligands A file or files containing the protein s or the part of a protein into which the ligand is to be docked This may be in PDB or MOL2 format see Setting Up the Protein s GOLD also needs a configuration file which contains the names of the protein and ligand files and all the user defined parameters such as genetic algorithm parameter settings fitness flags etc The configuration file can be created manually but it is usually easier and preferable
75. run the Diverse Solutions Stats are printed in the ligand log file e g 87 9 3 2 88 Move attempts 151617 Move failures 8964 Failure rate 0 059 e Asthe run progresses the failure rate will probably increase for each subsequent solution as it becomes increasingly more difficult to generate diversity Setting Up GOLD to Generate Diverse Solutions e To generate diverse solutions for a docking run click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window then enable the Generate diverse solutions check box e To specify the diversity criterion click on the Diverse Solution Options button The resulting Diverse solutions dialogue enables the following two criteria to e specified Cluster size the default is 1 Use this to specify how many ligand diverse solutions are contained in a cluster within a user defined RMSD R M S D the default is 1 5 A Use this setting to define the RMSD cut off in A for determining if diverse solutions are in the same cluster or not e The ligand log output file contains information on which ligands are in which cluster at a particular RMSD cutoff For example the Cluster size was 3 and the R M S D setting was 1 5 in the docking below Clustering method complete linkage Structure ids in cluster table rank nos Ordering of clusters and their members by rank order if from rms_analysis Distance Clusters 0 19 LT 2 es
76. terms or both To include weighted terms enable this check box Unweighted terms Enable this check box to include non weighted scoring function terms in the output file 119 13 3 120 Do not write SD style tags to Mol2 files Enable this check box to prevent SD style tags being written to comment blocks in MOL2 solution files Preserve COMMENT fields from input Mol2 ligand files Enable this check box to retain the COMMENT fields from input MOL2 ligand files in the docked solution files Save lone pairs Some 3rd party programs have difficulty reading files which contain lone pairs You can stop GOLD including lone pairs when it writes docked solution files by switching off this check box Save protein rotated atom positions Enable this check box to save the optimised positions of rotated protein atoms These include the optimised positions of polar protein hydrogen atoms and also final positions of any protein side chains that have been defined as being flexible Protein atom positions that are generated during docking will usually be different for each docked ligand pose and are therefore written to the individual ligand solution files Rotated atom positions are utilised by both Hermes and GoldMine Save per atom scores Enable this check box to include the scoring contributions of individual ligand and protein atoms to be written to docked solution output files For each atom its contribution to the total fitne
77. the constraint and the actual distance observed in the docked solution From your bestranking Ist file identify GOLD s top ranked solution for the ligand with the best total fitness score Go to the Hermes 3D view The overall top ranking ligand can be viewed by ordering the ligands based on their fitness score To do this go to the Molecule Explorer and find the tab labelled PLP Fitness Click on this tab either once or twice until you have the highest score i e the highest value listed at the top of the column The position and orientation of the terminal sulphonamide groups in the docked solutions should be similar to that observed in the co crystallised ETS inhibitor i e coordinated to the zinc within the protein via the sulphonamide nitrogen In the example below the terminal sulphonamide group of GOLD s top ranked solution can be seen to satisfy the specified constraint and reproduces the known binding mode of the co crystallised ETS inhibitor 177 178 This ends the tutorial GOLD User Guide 20 5 Tutorial 5 Docking with Water in the Binding Site Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial5 toa directory to which you have write permissions The object of this tutorial is to investigate docking to a binding site that contains water molecules which a ligand may either displace or alternatively make use of through hydrogen bond interactions The protein
78. the GOLD interface and read in the non_flexible conf file again The settings in this file are for the non flexible docking click on the Protein 1fax coagulation factor tab and click on the Flexible Sidechains option where we will be able to specify sidechains that will be allowed to rotate during docking Scroll down the list of residues until you get to GLN192 Click on GLN192 then hit the Edit button alternatively simply double click on GLN192 This launches a dialogue where we can add edit or delete rotamers GOLD User Guide Edit Rotamer Library GLN192 e The dialogue is blank as we have yet to define any rotamers for GLN192 e Click on the Library button Edit Rotamer Library GLN192 180 75 65 65 85 GOLD User Guide 189 The dialogue has been updated to reflect rotamers taken from a rotamer library in this case information taken from the paper The Penultimate Rotamer Library S C Lovell J M Word J S Richardson and D C Richardson Proteins 40 389 408 2000 It is a compilation of the most commonly observed side chain conformations for the naturally occurring amino acids The rotamer library txt file can be viewed in txt format in lt install dir gt goldsuite 5 3 GOLD gold Note that the library settings are simply a starting point users are encouraged to generate their own rotamers for optimal results The nine allowed rotamers are listed at the bottom of the Edit Rotamer Library
79. the check box labelled Generate a cavity atoms file from the selection By enabling this option the binding site definition will automatically be expanded to include all atoms in the existing definition plus all the atoms of their associated residues To manually refine this selection click on the Refine Selection button to open the Refine Binding Site Selection dialogue All residues included in the binding site definition are listed Residues can then be added or removed from the selection by clicking on atoms in the Hermes visualiser The cavity atom selection can be saved as a protein atom subset and viewed within Hermes To do this click on the Add Definition as a Selection button You can then highlight the atoms belonging to the subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area Note that it is not possible to define the binding site from an atom when performing an ensemble docking Defining a Binding Site from a Point Click on Define Binding Site from the list of Global Options given on the left of the GOLD Setup window Switch on the button labelled Point Then within the Hermes visualiser click on one or more protein atoms in order to define a centroid close to the centre of the active site Alternatively the orthogonal x y z coordinates of a single solvent accessible point approximately at the centre of the active site can be typed directly into the t
80. the column labelled Type Next specify the protein atom that forms the interaction This can be done by clicking on an atom in the visualiser Alternatively you can enter the residue chain name and atom identifiers directly into the appropriate entry boxes GOLD User Guide GOLD User Guide Note that your protein input file must include chain identifiers in order avoid problems when specifying atom for use in this constraint A maximum of 10 interactions can be defined Note that hydrogen bond acceptors that can form either a hydrogen bond or a weak CH O interaction need to be added twice one for each type of interaction To delete an interaction click on the corresponding row number to highlight the interaction definition then hit the Delete Interaction button One or more binding motifs can now be defined Each motif should consist of a unique combination of the interactions that you previously specified To define a motif Click the Add Motif button Specify each of the interactions that need to be included in the motif The hydrogen bond interaction types Acceptor Donor CHO are set as either 1 or O depending on whether or not they are observed in a particular motif The lipophilic interaction type is set as the frequency of that interaction as observed in the set of complexes used to originally identify the motifs The frequency of a lipophilic interaction is added to all the interaction motifs To delete a motif c
81. the internal energy and clash coefficients respectively C is per default set 0 2 and both Cin and Ccigsh are Set to 1 0 To speed up scoring and docking grids are precalculated for each atom type using a grid spacing of 0 3 A The max distance for interaction 6 0 A the scaling factor C and the respective weights for the internal Cint and clash energy Caasn together with the grid spacing can be altered in the asp params file see Altering ASP Fitness Function Parameters the asp params File 7 5 4 Metal and Hydrogen Bond Correction e In ASP you can add a correction when docking to metal containing receptors When adding the metal correction to the ASP score a hydrogen bond correction is included by default The hydrogen bond correction is similar to the one found in ChemScore see Hydrogen Bond Terms The final score is calculated as ASP Fitness Col S map S hbond S metal Cnt pint clash clash e The S metal is calculated for single metal ligand atom interactions based on the actual distance between the metal and the ligand acceptor and is corrected for the metal ligand score in S map This means that the S map contribution based on grid points is subtracted from S metal to offset the contribution that is already present in the ASP grid for the acceptor type The S metal score more accurately reflects the metal ligand interaction e The S hbond correction corresponds to the metal correction as it calcu
82. the protein s can now be used with GOLD but only if the modifications we have made are saved out as a MOL2 file Cavity atoms this option will be greyed out e Ensure the GOLD conf file and Protein tickboxes are activated and that the filenames are as you want them then hit Save to start the docking Things to Consider While the Docking is Running e Protein and ligand initialisation Both the protein and ligand files are initialised before the docking commences At this step GOLD deduces atom types from the information about element types and bond orders in the input structure files It is therefore crucial that both the protein and ligand input files are prepared according to the guidelines provided see Atom and Bond Type Overview 152 GOLD User Guide If automatic atom typing is switched on GOLD will re type any atoms or bonds it considers to be incorrect and will issue a warning concerning this in the gold err file However if for any reason GOLD is unable to deduce an atom or bond type then the atom or bond in question will be replaced with a dummy atom type Du or an unknown bond type Un respectively By default automatic atom typing is carried out on the ligand file but not the protein file however this can be enabled under the Atom Typing Advanced option e Protein flexibility By default the torsion angles of Ser Thr and Tyr hydroxyl groups will be allowed to rotate during docking in order to optimise their hyd
83. the scores of the function are close to zero at this length The two terms f and f denote the protein and ligand volume corrections to the contacts respectively These two terms are added to account for the difference in accessibility of different protein and ligand atoms if no excluded volume corrections are included in the reference state the expected number of contacts is simply the product of the average contact density in the sphere with radius Rmax and the volume of a spherical shell at distance r The way that these corrections are defined differs from other potential scoring functions and the inclusion of a protein correction term is novel to the ASP fitness function The ligand correction term can be compared to the corresponding term included in Drugscore The Generation of Potentials To derive the pair potentials between ligand and protein atoms a database of protein ligand complexes from the PDB is used Bond types of the protein are assigned based on residue and atom names Additional atom types are defined to separate backbone nitrogen and oxygen atoms from those in aspargine glutamine aspartic acid and glutamic acid side chains and to distinguish serine and threonine hydroxyl oxygen from those in tyrosine Ligands in the database were divided into three separate categories i covalent ii co factor and iii normal Only binding sites with normal ligands with a heavy atom count between 6 and 60 were included in the database co fac
84. to vary ligand ring conformations during docking The use of ring conformation templates may improve the chances of GOLD finding the correct answer by allowing the algorithm to sample ring conformations that are commonly observed in crystal structures Rings in the ligand are matched against a template library If a matching template is found then the conformation of that ligand ring will be varied during docking using a set of supplied alternative conformations for that ring Each time an alternative conformation is sampled the ligand conformation is changed to match the new conformation by altering the bond lengths internal ring angles and torsions A ring torsional strain energy term is also added to the ligand internal energy To use ring conformation templates during docking click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window activate the Match template conformations tick box in the Explore ring conformations section of the window Information on the composition of the CSD ring conformation library and how ring templates are matched at run time is available see The CSD Ring Conformation Library and Matching Templates at Run Time It is also possible to specify your own ring templates and the allowed alternative conformations for those rings see User Defined Ring Conformations 8 1 3 The CSD Ring Conformation Library and Matching Templates at Run Time GOLD User Guide GOLD id
85. tutorial into the GOLD front end From the Global Options on the left hand side of the GOLD front end click on Output Options Specify or browse to a directory using the button adjacent to Output Directory for which you have write permission This is where the GOLD output files will be written GOLD User Guide The Handling and Parameterisation of Metals in GOLD e GOLD is able to predict binding to seven metal ions Mg Zn Fe Mn Ca Co and Gd e No special instructions are needed to dock to metal ions they will be handled automatically when present in the protein binding site Automatic Determination of Metal Coordination Geometries e GOLD will automatically recognise the following metal coordination geometries Template Geometry Coordination Number TETR Tetrahedral n 4 TBP Trigonal bipyramidal n 5 OCT Octahedral n 6 CTP Capped trigonal prism n 7 PBP Pentagonal bipyramidal n 7 SQAP Square prism n 8 ICO Icosahedral n 10 DOD Dodecahedral n 12 e In order to determine the coordination geometry of a particular metal atom GOLD performs a permuted superimposition of coordination geometry templates onto the coordinating atoms found in the protein Coordination fitting points are then generated using the template that gives the best fit based on RMSd e The geometry templates used for a given metal are defined in the gold params file in the section headed Metals H Bonding Sybyl atom ae Le pDanor D Allowed 7 Coord
86. weight previously specified see General Methodology The details of each specified protein H bond constraint satisfied in the solution are listed and an overall constraint score is given A list of all hydrogen bonds formed between ligand and protein is also provided in the ligand log file Go to the end of the gold_ligand_m1 log file then scroll up slightly until you see text similar to the following Final ranked order of GA solutions BZ ih This text tells you how the docking attempts rank in terms of fitness score So the third docking attempt is the top ranked while the first docking attempt is the lowest ranked of all the solutions Go to Hermes 3D view and display the top ranked solution note that it may not be docking attempt number 3 If you are still unsure which is the top ranked solution the docking results can be ordered based on their fitness score in the Molecule Explorer window using the PLP Fitness header in the Docking Solutions view Inspect how well the docked inhibitor fits within the protein binding site as predicted by GOLD GOLD User Guide e Interactions between the cyclic urea inhibitor and HIV 1 protease can be divided into two groups those that anchor the scaffold in the active site and those that fix the substituents in the target subsites e Confirm that the hydrogen bonds specified in the constraints are formed as expected to the cyclic urea scaffold by measuring the relevant contact dist
87. worst fitness The numbers in the matrix of rms deviations refer to the rankings not the run numbers e g row 1 of the above matrix refers to the solution with the best fitness score contained in ranked_ligand_m _1 mol2 Finally the rms deviations are used as input to a hierarchical cluster analysis using the complete linkage algorithm Each line shows one iteration of the clustering algorithm the distance between the clusters that were merged at that step and the contents of the current set of clusters Clusters are separated by the symbol and rankings are used rather than run numbers For example the solutions ranked_ligand_m 2 mo12 and ranked_ligand_m 4 mol12 were merged in the first step of the following cluster analysis Clustering using complete linkage Structure ids are RANKING Dist Clusters 3 14 4 06 5 07 10 95 4 2 3 5 1 4 2 3 1 5 4 2 3 1 5 4 2 3 1 5 Identification of Different Binding Modes Clustering of Ligand Poses GOLD clusters docked solutions according to how similar the poses are in terms of their RMSd see Comparison of Docking Solutions A link can be generated to the top ranked solution from each distinct cluster This can be useful in identifying different ligand binding modes Considering solutions from different clusters is often more relevant than taking the top n ranked poses since these will often be very similar i e all from the same cluster of sol
88. 0 4 1 Method Used for Region Hydrophobic Constraints e This constraint can be used to bias the docking towards solutions in which particular regions of the binding site are occupied by specific ligand atoms or types of ligand atom e For each region hydrophobic constraint specified a sphere is placed at an explicitly defined position within the binding site Each sphere is assigned a user defined radius so a sphere can be adjusted if required e g to fill an entire pocket in the binding site Minimum settable radius as 0 5 A e Acontribution determined according to a user specified weighting is then added to the score for each specified non hydrogen ligand atom that lies within the designated sphere A contribution is added to the score for each atom located within the sphere i e the total contribution will depend on the number of atoms found in the region of interest and ultimately the ligand accessible volume of the region e The ligand atoms used in the constraint can be specified explicitly Alternatively it is possible to use all hydrophobic ligand atoms or to use only those hydrophobic atoms in aromatic rings Atoms considered to be hydrophobic include Carbon atoms bound to at least two H or C atoms Atoms typed C cat Atoms typed S 3 and bound to two carbons _Hatoms bound to an sp2 sp3 or aromatic carbon Note Only heavy atoms found within the sphere will contribute to the score e Details of the region
89. 0 GOLD User Guide Protein Setup GOLD User Guide W GOLD Setup EF Wizard step 2 Protein setup At this point you have the chance to edit your protein structure if required e g add hydrogens delete waters Global Options 1TBF 1t9s ixp0 2chm Wizard steps To edit the protein s use the options available on the protein tabs to 1 Select a protein 1 Add Hydrogens 2 Protein setup R 3 Define the binding site 2 Configure active waters and delete unnecessary waters 4 Configuration template 5 Select ligands 3 Delete ligands 6 Choose a fitness function 7 GA search options 8 Finish Help R lt Back Next gt Cancel Wizard In the Hermes 3D view you will notice that three of the proteins 1t9s 1xp0 2chm are superimposed and have had hydrogen atoms added The fourth protein 1tbf has not been prepared it has no H atoms is in a different frame of reference to the other proteins and still contains ligands An essential step of protein set up for ensemble docking is that the proteins are superimposed This is because there should be only one binding site definition across the whole ensemble In the Hermes 3D view you will notice the Molecule Explorer off to the left hand side of the Hermes interface Click on the adjacent to 1TBF and underneath All Entries Right click on Ligands and select Delete from the pull down menu Return to the GOLD wizard and click on the 1TBF tab adjacent to the Global Options tab
90. 0075 3 8074 13 2320 c 3 1 RES1 0 0000 ates 6 7642 4 9497 12 6823 C 3 1 RES1 0 0000 6 2 3 6 5491 5 0625 11 4378 C 3 1 RES1 0 0000 lt TRIPOS gt BOND 1 1 2 1 2 3 2 1 3 4 3 1 4 5 4 1 5 6 1 1 6 5 6 zl e Secondly a set of allowed alternative ring conformations for each ring in template _library mol2 must be created and stored in the following files lt GOLD DIR gt user ring _conformations 0010001 mo12 lt GOLD DIR gt user ring _conformations 0010002 mo12 and so on The ring conformation filenames are based on the corresponding template identifier In the above example the triazole ring template has the molecule identifier 0010001 The alternative conformations of that ring that will be used during docking must therefore be located in lt GOLD_DIR gt user_ring_conformations 0010001 mol2 e Note that any user defined templates and their corresponding set of allowed conformations will be used in preference to the supplied CSD conformations e Note User defined ring conformation functionality does not extend to fused rings or macrocycles A work around would be to create a separate template that matches the ring of interest in the fused ring however this may give rise to strange results in some instances thus should be used with care GOLD User Guide 79 8 2 Flipping Amide Bonds During ligand initialisation ligand amide groups including thioamides ureas and thioureas will be set to the trans conformation Flattening t
91. 12 e In order to determine the coordination geometry of a particular metal atom GOLD performs a permuted superimposition of coordination geometry templates onto the coordinating atoms found in the protein e g if there are only two coordinating atoms in the protein then every unique pair of coordinating template atoms are selected and superimposed on the system in the protein e Coordination fitting points are then generated using the template that gives the best fit based on RMSD e The geometry templates used for given metals are defined in the gold params file in the section headed Metals for explanation of parameters refer to comments in the gold params file Atom Type Donor D Allowed noig Sale default or Acceptor A Coordination Coordination Type Type elucidated or Metal M Geometries Distance MGD Mg DEF M 4 6 2 05 ZND Zn DEF M 4 5 6 2 09 MND Mn DEF M 4 6 2 06 FED Fe DEF M 4 6 1 98 CAD Ca DEF M 6 7 2 44 COBD Co oh DEF M 6 2 09 GDD Gd DEF M 6 2 44 e For example for a Zn atom GOLD will attempt to match coordination geometries 4 5 and 6 tetrahedral trigonal bipyramidal and octahedral templates onto the coordinating atoms found in the protein e The template that gives the best match will then be used to generate coordination fitting points e Details of the coordination geometry determination are given in the gold_protein log file e The output file gold_protein mol2 will contain a number of dummy ato
92. 258 Both moieties have been highlighted in capped stick in the image below note that the H atoms on the protein have been hidden for clarity GOLD User Guide 203 If you view the protein and ligand files separately toggling either off using the Molecule Explorer you will notice the N atom is present in both files The presence of this atom in both files is extremely important as will be illustrated when we set up the docking later on Close the protein and ligand files by clicking on File and then Close All Files in the Hermes main menu Setting up a covalent docking 204 Open Hermes Load the gold conf provided into GOLD via GOLD Setup and Runa Docking Load Existing then navigate to the folder containing the tutorial9 files select gold conf and click Open Open the ligand mol2 file via File Open The settings for a covalent docking are specific to the protein so click on the Protein lase aminotransferase tab to access the protein settings then click on Covalent Activate the Define covalent docking tickbox As we are docking a single ligand and not a substructure ensure the Ligand link mode Atom radio button is selected Now we must either enter the atom IDs manually into the Protein link atom and Ligand link atom boxes or else select the atoms in the 3D view Both the ligand and protein files are open in the 3D view so return to Hermes and select first the ligand link N atom atom ID 25 and the protein link atom
93. 3 ribose C 3 ribose N am uracil C 2 1H C 2 uracil O 2 24 73 85 44 59 60 40 14 83 2000000000000075 300 143 3510 6 DIAGRAM benzyl sub C C 3 2H C ar C ar OH C ar OH expand 0 0 180 0 Oo000000000000000000000 0 1 9 27 76 64 15 7 420000 GOLD User Guide 235 25 4 Extracting Torsion Angle Distributions from the Cambridge Structural Database The command process_tab only available on SG machines will extract the torsion angle histogram from the tab file produced by a search of the Cambridge Structural Database and reformat it so that it can be added into the GOLD torsional distribution file 236 GOLD User Guide
94. 4 30 30 30 32 na 27 04 87 82 24 61 09 13 55 86 79 79 93 91 60 80 34 ra 28 28 27 27 28 27 73 oo 59 85 04 41 23 00 14 29 29 29 76 40 78 45 42 an Sihb_int 00 gold_ligand_m1 log Siint 29 11 14 8 15 29 15 04 Interrupt G amp f view Solutions Close of population s 3 size 100 selection pressure 1 100000 58 19 04 89 22 58 22 22 82 82 Ard Bord 82 84 84 na Sicov azz z0 F 21 F 13 7 ixi Scroll through the docking results You will notice the ligand is indeed bonded to the protein The docked poses can be compared to the pose of the native ligand structure by superimposing the docking solutions with the ligand mol2 file Select the Display tab at the top of the Molecule Explorer and ensure only the Ligand_reference 1ase aminotransferase tickbox is active then return to the Docking Solutions tab and select all the docking solutions 207 This ends the tutorial 208 GOLD User Guide 20 10 Tutorial 10 Ensemble Docking Introduction Recently it has become less a case of Is there a model available for my target and more a case of How do I make use of all the structural data available for my target One way of addressing this is to dock into multiple protein models i e ensemble docking Ensemble docking is also a way of modelling
95. 50 and a Maximum separation of 2 50 distances are in A As with standard distance constraints the fitness score is reduced for solutions which do not satisfy the constraint The amount by which the score is reduced is determined by a user defined weight term Set the value of the Spring constant to 20 0 then click on the Add button to add the constraint to the constraints list 175 3 GOLD Setup SBE Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial4 gold conf Load Save Options Protein 1cil lyase oxo acid Protonation amp Tautomers Waters Protein atom number 2041 Delete Ligands miexi sarehan Substructure file C Documents and Settings henderson Desktop gold_tutorials tutorial4 substructure mol2 oft Potentials Metals 5 Constraints Substructure atom no 4 Distance P Use ring centre nearest to selected atom ring atoms only Substructure HBond Minimum separation 15 Protein HBond Covalent Maximum separation 2 5 Interaction Motif Spring constant 5 0 Reset Details Substructure protein 2041 C Documents and Settings henderson Desktop gold_tutorials tutorial4 substructure mol2 4 2 Help R Run GOLD Run GOLD In The Background Finish Cancel e Click on the Global Options tab to return to the general docking setup window Running GOLD e The time taken by GOLD to dock ligands can be controlled by altering the values of the genetic algorit
96. 9 4 1 5 30 An Energy may be assigned to a given rotamer This will penalise i e reduce the fitness by the value specified if the side chain is placed in defined conformation In other words it makes this conformation ess favourable A negative Energy value can be entered its effect would be to improve i e increase the fitness if the side chain is placed in the defined conformation Allowing a Localised Backbone Movement Quite often a side chain rotation is accompanied by a small change in the local backbone conformation For example the figure below shows a detail from an overlay of two PDB structures 1qon 1dx4 of the same enzyme Not only has the Tyr side chain rotated around Ca CB and CB Cy but there has also been a small backbone movement primarily affecting the position of the Ca atom Although minor the two Ca positions are only 0 6A apart this movement is extremely important because it alters the vector direction Ca CB and this can have a big leverage effect on the positions of atoms further down the side chain In this case it is impossible to overlay the Tyr370 side chain of 1dx4 closely onto that of 1qon simply by rotating around the Ca CB and CB Cy bonds This is about as close as one can get GOLD User Guide GOLD User Guide The backbone movement can be mimicked by allowing the Ca atom and the attached side chain to rotate around the N C vector where N and C are the backbone atoms on either side
97. Apply localised soft potentials to specific residues Add selections Clear Residues alternative potential 2 GLN192 Add selections TYR99 Clear Click on atoms in the visualiser to add them to the above lists Clicking an atom For a second time will remove it from a list Help R Run GOLD Run GOLD In The Background Finish Cancel Select the alternative potential you wish to apply by switching on the corresponding Add Selection button then specify those residues you want to apply the alternative potential to by clicking on them in the Hermes visualiser Selected residues will be listed GOLD User Guide To remove a selected residue from the list click on it again within the Hermes visualiser To remove all selected waters click on the Clear button More than one residue can be specified and both alternative potential forms can be used in the same GOLD run as shown in the above example GOLD User Guide 39 5 5 1 5 2 40 Setting Up Ligands Essential Steps Add all hydrogen atoms including those necessary to define the correct ionisation and tautomeric states see Ligand Hydrogen Atoms lonisation States and Tautomeric States Ensure that all bond types are correct If they are and hydrogen atoms have been placed on the correct atoms GOLD will deduce atom types automatically when atom typing is turned on see Automatically Setting Atom and Bond Types GOLD assigns at
98. D setup wizard by clicking on the main menu option GOLD then by picking Wizard from the resultant pull down menu The steps required to set up files for docking are listed down the left hand side 209 n GOLD Setup Conf file Load Save Global Options GOLD has many configuration options which are available via the tree view at the left of this Templates dialog To simplify the process you can dick the Wizard button below and you will be guided Proteins through the essential configuration steps At the end of the wizard you can either start the Define Binding Site GOLD docking or access the more advanced configuration options Select Ligands Configure Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing Help R RunGOLD Run GOLD In The Background Finish Cancel e Inthe Select one or more proteins dialogue read in the four protein files 1t9s mol2 1tbf pdb 1xp0 mo1l12 and 2chm mo12 one at a time by hitting the Load Protein button navigating to the folder to where you copied the tutorial10 files select each protein file and then click on Open Please note that whilst there is a maximum limit of 20 proteins when using ensemble docking we do not recommend using more than 10 proteins e Note that as each protein is added a tab corresponding to that protein appears to the right of the Global Options tab 21
99. DB information can be taken from a smaller set of crystal structures which could be comprised of only one family of proteins or if there is insufficient information in the specific target database one can mix in information from the general PDB database e You can store your customised potentials in a directory specified in the asp params file see Altering ASP Fitness Function Parameters the asp params File 7 5 7 Performance of the ASP fitness function e Onthe CCDC Astex validation set ASP has similar success rate as Goldscore and Chemscore for a more complete discussion on the accuracy of the ASP please refer to the original publication see Overview 7 5 8 Altering ASP Fitness Function Parameters the asp params File e The ASP parameter file is stored in the GOLD_HOME gold directory It contains all the parameters used by the GOLD implementation of ASP A full description of the meaning of the ASP specific parameters is given below e The ASP file can be customised by copying it editing the copy and instructing GOLD to use the edited file e To use a modified asp params file click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select GoldScore from the Scoring Function drop down menu Then either enter the path and filename of the Scoring function parameter file or click on the button and use the file selection window to choose the file e The format of th
100. DED HAL S During docking GOLD selects a list of lipophilic ligand atoms and matches them onto a subset of the hydrophobic fitting points GOLD User Guide It is possible to use customised hydrophobic fitting points This might be appropriate if GOLD is not giving good results on a particular protein and you suspect that the fault may lie in the placement of hydrophobic ligand groups Customised fitting points must be supplied in a MOL2 format file that contains a list of dummy atoms at the desired fitting point locations The supplied fitting points should sample all regions of interest in the cavity so that the docking algorithm has sufficient alternatives for placement of hydrophobic ligand atoms within the cavity GOLD uses gridded points that are spaced by 0 25 A for a speed up in calculation higher values could be used To make GOLD use a customised fitting point file click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and enable the Read hydrophobic fitting points check box Then either enter the path and filename of the MOL2 file or click on the button and use the file selection window to choose the file Customised fitting points can for example be generated by the CCDC program SuperStar which offers the possibility of writing out a file of GOLD fitting points in the appropriate format see SuperStar manual sections on SAVE_GOLD_FITTING_P
101. Display tab in the Molecule Explorer then right click on the reference ligand From the resultant menu it is possible to modify the representation style and colours of the reference ligand You may also wish to GOLD User Guide simplify the display by hiding the protein H atoms using the Hide H button at the top of the Hermes interface or by suppressing the protein altogether by deactivating the Protein 1acj hydrolase carboxylic esterase tickbox Analysis of results All waters turned off From Load GOLD results in Hermes read in the gold conf file corresponding to your second set of results waters_off conf The second set of results are read in after the first set allowing you to directly compare docking results from different runs If it is not clear which solutions correspond to which docking run expand the Docking Solutions header at the top of the ligand solutions list Docking Solutions Display Movable Customise Sor Group by No grouping 7 Colours Deci Solutions Chen ligand from 1ACJ soln 1 24 9566 tacrine igand from 1ACJ soln 2 235046 tacrine ligand from 1ACJ soln 3 27 3380 tacrine ligand from 1ACJ soln 4 24 9279 tacrine ligand from 1ACJ saln 5 24 4033 tacrine ligand from 1ACJ saln 6 25 0178 tacrine ligand from 1ACJ soln 25 5124 tacrine ligand from 1ACJ soln 8 24 4341 tacrine ligand from 1ACJ saln 9 24 4861 tacrine ligand from 1ACJ saln 10 25 2309 tacrine ligand fram
102. F SYB_TYPE EL_TYPE C 3 C 2 C 1 C ar Cat N 3 N 2 N 1 Near N am N pl3 N 4 0 3 0 2 O co2 S 3 S 2 S o S 02 P 3 H FI C Br CIINIIO ISIIP e This grammar allows torsions to be specified as four fragment nodes Each node defines an atom type and optionally a set of neighbours to which the atom is connected Each of the neighbours is a node or an exact count of the number of hydrogen atoms to which the atom is bonded Atom types are defined using SYBYL atom types or elemental atom types The atom can also be required to be part of a pre defined fragment e Bonding environments can also be specified using the symbols which indicate respectively that an atom forms an aromatic double or single bond to its parent node Note and should therefore not be used on the first atoms specified these bond types are specified for substituents only e Anode isa parent of all its neighbours and a top level node in the torsion definition is a parent of subsequent nodes in the torsion e There are currently four fragments available one of which the uracil fragment matches both thymine and uracil More fragments can easily be added The Ullman algorithm is used to determine if an atom belongs to a fragment Fragments are defined through SYBYL atom types and connectivity exact bond types are not used Only heavy atoms are considered Currently fragments are precompiled
103. File which Contains a Metal lon 20 3 9 2 Automatic Determination of Metal Coordination Geometries 20 3 9 3 Specifying Metal Coordination Geometries Manually c00008 22 3 9 4 Defining Custom Metal Coordination Geometries cccccesseeeeees 23 3 9 5 Metal Ligand Interactions ccccccccccccccseessseeceeceesseeeseeseceesessaaanees 24 3 9 6 Heme Containing Proteins ccccccecccccesssececeeeseeeeeeeeeeeseeeseesaeeseeees 24 4 Protein FI XiDIity ccccccesesseeeccccecceeeeseeececeeeesaeesseeeeeeeeessaeaesseeeeeeeessaaeaseeeeeesessaaagaass 25 4 1 Side Chain Flexibility cccceccctetcnceevenscetctetanctetetvecys teense vee cea tate see oe ea See 25 4 1 1 aide s e b letd eo peeeeerer reese rree ere reece cree reer rreeerer er EEEE EE reece reer 25 4 1 2 Specifying Flexible Side Chains cccsssccccccecceesseseeeeeeessaeaeseeeeeess 25 GOLD User Guide 4 1 3 Defining ROtAMESS ccccccccccssssseseecccceecaeeesseeceeeeesseaaseeseeeeeessaaaness 27 4 1 4 Deleting and Editing Rotamer Definitions cccccssssssseeeeeeseeeeees 29 4 1 5 Allowing a Localised Backbone MoveMe nb ccccccccsssseeeeeeeeeeeeees 30 4 1 6 Protein Protein ClaShes eccessscceeeeeeeeeeessceeeeeeeeeeeseaaaeeeeeseeeeees 32 4 2 Large Backbone Movements c cscscccccccsseesssesecceecesaaesseeeceeeeessaaaasseeeeeeeesaaa 33 4 3 Ensemble DOcking cccccccc
104. GOLD User Guide A Component of the GOLD Suite 5 3 Release Copyright 2014 Cambridge Crystallographic Data Centre Registered Charity No 800579 Conditions of Use The GOLD suite of programs the Program comprising all or some of the following Hermes including as Relibase client and as SuperStar interface GOLD GoldMine associated documentation and software are copyright works of CCDC Software Limited and its licensors and all rights are protected Use of the Program is permitted solely in accordance with a valid Software Licence Agreement or a valid Licence and Support Agreement with CCDC Software Limited or a valid Licence of Access to the CSD System with CCDC and the Program is proprietary All persons accessing the Program should make themselves aware of the conditions contained in the Software Licence Agreement or Licence and Support Agreement or Licence of Access Agreement In particular e The Program is to be treated as confidential and may NOT be disclosed or re distributed in any form in whole or in part to any third party e No representations warranties or liabilities are expressed or implied in the supply of the Program by CCDC Software Ltd its servants or agents except where such exclusion or limitation is prohibited void or unenforceable under governing law GOLD 2015 CCDC Software Ltd Hermes 2015 CCDC Software Ltd GoldMine 2015 CCDC Software Ltd Implementation of ChemScore Heme Kina
105. GOLD gold_soln_ligand_m3_8 mo12 LIN NTS_262_ pdbicin_1 48 69 14 41 31 76 0 00 9 38 home GOLD gold_soln_ligand_m4 9 mo12 LIM INQ_555_pdbii9i_1 46 35 14 08 31 06 0 00 10 44 home GOLD gold_soln_ligand_mS_2 mo12 LIM INM_555_pdb1i90_1 15 1 6 Rescore Solution File e A file containing the docked ligand solution s after rescoring can be written You can control whether or not this file is written see Rescore Output Files e If specified solutions will be written with the default filename rescore mol2 To specify an alternative filename for both the rescore solution and log files add the 128 GOLD User Guide following line to the gold conf file concatenated output lt filename mol2 gt For example if concatenated output Myfile mol2 the rescore mol file will be named Myfile mol2 Solution files will contain the new scoring function terms and the positions of rotatable protein hydrogen atoms generated during rescoring see Rescore settings A full description of the additional tags written to solution output files is available in see Appendix C Additional Tags in Output Files 15 1 7 Rescore Log File The rescore log file rescore 1og summarises the outcome of the rescoring run To specify an alternative filename for both the rescore solution and log files add the following line to the gold conf file concatenated output lt filename mol2 gt For example if concatenated output Myfile m
106. Genetic Algorithm Parameter Settings 113 12 R SCOFINE encarta er a ewe ees ee Ae ee E E 114 LA AWENN ear 25 be5 5 Sacco co weno een sere E T N T ene en tere enone Benmore enero meant 114 12 2 Setting Up a Rescoring RUN cceccccesseececcaeesceeneeeeceeaeeseeessaseessueeeesaeeeeeees 114 12 3 RESCONE SCTLINES eet eee eee ee a ee ee ee ee ee 115 12 4 Receptor Depth Saline rescence ees ee ee EE 116 12 57 Res ore OUtpUt Files SA ce Se 2Y Bes SETA OS ORI E h a aE a i 117 13 Docked Ligand Output Options ossein i ait i aaa 118 13 1 Specifying Ligand Solution File Formats and Directories ccccccceeeseeeeees 118 13 2 Controlling the Information Written to Ligand Solution Files c00eee 119 13 3 Selecting Which Ligand Solutions to Keep sessssssssssnesssssssrennrssssssrerreessssse 120 14 Running GOLD Aerian eee enrere etua eoulen sees oastaSuscdcucgesctctssesatotesseatovesseatoseseeatosesstitseesexs 122 14 1 Required Input Files c i ccccceeccccccccseeesseeeceeeeseaeeeseeeceeeesseaeesseeeeeeeessaageess 122 14 2 Running GOLD Interactively cn008 cc awd oad ei ek aa edd eek eee es 122 14 3 Submitting a GOLD job to the Background ccceesescecccceeceeeeeeeeeeeeessaaaaeess 124 14 4 Running GOLD from the Command Line ccceeseeeeecccecceeeeeeeeeceeeeeeaeaaness 124 14 5 Running in Parallel ccccccsssssscccccecceeesseeecceeeeseeeeeseeee
107. LD User Guide As with the protein file all hydrogen atoms must be present in the ligand input file We have already added H atoms to the ligand 1xoz_ligand Specify the ligand by hitting the Add button at the bottom of the GOLD Wizard Navigate to the folder to which you copied the tutorial10 files select 1xoz mo12 then click Open The 1x0z mol2 is now listed under Ligand File The number of dockings to be performed on each ligand is specified under GA runs by default this value is 10 The value can be edited by clicking in this window and re entering another value Increase the number of times the ligand is docked to 20 GA runs 213 GOLD Setup a Wizard step 5 Select ligands Choose one or more ligands to be docked into the protein by clicking the Add button Global Options 1TBF 1t9s 1xp0 2chm Wizard steps z z 1 Selecta protein Ligand File GARuns FirstLiqand Last Ligand 2 Protein setup 1 ixoz mol2 20 llast 3 Define the binding site 4 Configuration template 5 Select ligands 6 Choose a fitness function 7 GA search options 8 Finish Show full file paths Add Delete Reference ligand Help R lt Back Next gt Cancel Wizard e Click Next to proceed to the Choose a fitness function dialogue Selecting a Fitness Function e During a docking run the solutions found by GOLD are scored according to a fitness function Ensure that the default CHEMPLP scoring functio
108. MF and Drugscore ASP has comparable accuracy to the ChemScore and GoldScore fitness functions Traditional scoring functions are based on force fields or on regression where parameters are derived from a set of experimental binding affinities and structures ASP uses a different approach information about the frequency of interaction between ligand and protein atoms is gathered by analysing existing ligand protein structures in the PDB and this information is used to generate statistical potentials Depending on the database where the atom atom potentials are taken from the scoring function created can be targeted to certain proteins see Targeted Scoring Functions A general scoring function one that can be used for all types of proteins would take its interactions from the entire PDB while a more targeted function would be created from specific families of proteins Atom atom potentials for a general scoring function are included in the GOLD distribution Empirical parameters used in the fitness function hydrogen bond energies atom radii and polarisabilities torsion potentials hydrogen bond directionalities etc are taken from the GOLD parameter file These parameters are independent of the scoring function being used Parameters can be customised by copying the file editing the copy and instructing GOLD to use the edited file see Altering GOLD Parameters the gold params File A scoring function specific parameters file
109. NE tOr E TEETER TEREKET OEE EEEE TA EENE NEKONA AITTA EEA 59 7 3 2 Van der Waals and Hydrogen Bonding Annealing Parameters 60 GOLD User Guide 7 3 3 Altering GoldScore Fitness Function Parameters the GoldScore Parameters File is ccccccccceccecscccscsacecescaesdiadsaescsuseceassesceedcceactancussceeaesenize 60 7 4 CHEMS CONCH 2 5 an erred ian E r R O E O E ET E E O E O E O sae 60 7 4 1 OQVEMIEW 11428888 RAHA E E eE U O eE E iE AH 60 7 4 2 Block Functions in CHEMSCOLE cccceccccccccccceeeeeeeeeeeceeeeeeeeeeeeeeeeeeeess 61 7 4 3 Hydrogen Bond Terms ccccccccssssssseeceeceecaeeesseeceeeeeeaeaaseseseeeeessaaaaess 63 7 4 4 Metal Binding and Lipophilic Terms ccccccccceeesssseeeeeeeeeeeeaaeees 65 7 4 5 Rotatable Bond Freezing Term ccccccccssssssseeccceeccaeeesseeeeeeeesseaanees 66 7 4 6 Clash Penalty and Internal Torsion Terms sssssseecceeeesssesseeseeees 67 7 4 7 Covalent Term oo cccccceeeeeeeeeeee eee a aa aa aaa aaa a aaa a aaa eee 68 7 4 8 Constraint Ter Sue ie eeo ae eei a EE E E EA E a i waist 68 7 4 9 Altering ChemScore Fitness Function Parameters the ChemScore File EE EE EEE E E EEEE E A EEE E E E 68 7 5 Astex Statistical Potential ASP cccccccccccceeeeeeeeeeeeeeeeeeeeeeeeeeeeeseeeeeeeeeeeeeeees 69 7 5 1 OAA ETE 69 7 5 2 The Reference State errs a TaT ETE ETERRA 69 7 5 3 The Generation of Potentials ccccccccccccccccccceeeeeeeeeeeeeeeeeeeeeeeeeeess 70 7 5 4 Metal and Hy
110. OINTS and GOLD MIN PROPENSITY 9 3 Generating Diverse Solutions There are occasions when GOLD obtains a number of docking solutions for a particular ligand which are very similar or if the Allow early termination option is activated GOLD may obtain a user defined number of ligands within the allowed RMSD very quickly Although this may not always be a problem there are occasions where it is apparent that none of the solutions are correct If this happens GOLD can be set up so that a number of different diverse solutions can be generated 9 3 1 Method Used to Generate Diverse Solutions GOLD User Guide Diversity is enforced during the ligand mapping stage As the ligand is constructed and mapped into the binding site the GOLD checks the RMSD of the current solution against those that have already been generated If the RMSD is below the diversity threshold or the maximum number of solutions per cluster has been reached the mapping is rejected and the process repeated until an acceptable solution is generated GOLD keeps track of any failures once the failure threshold has been reached the diverse solutions code is switched off The failure threshold is checked once the diverse solutions code has been called a thousand times After that if the ratio of the number of failures to the number of times the code is called i e the number of attempts is greater than 0 2 then the diverse solutions code is switched off After each GA
111. Output e Click on the Run GOLD button at the bottom of the GOLD front end You will be prompted that the GOLD configuration has been updated and needs to be saved click Save to proceed to the Finish GOLD Configuration window 3 Finish GOLD Configuration 21 x Directory foaram Files CCDC GOLD Suite GOLD examples tutorial2 ey Save Files JV GOLD conf file gold conf Protein 1442 lyase_protein mol2 Tl Protein s ie Cavity atoms cavity atoms ms e From within this window we can specify the directory the gold conf is to be saved to we will leave this as the default working directory save a protein mol2 file and specify its name this is only necessary if the protein file has been modified We have not modified the protein mol2 file so ensure the Protein tickbox is deactivated save a GOLD conf file and specify its name We have changed some of the configuration options so ensure the GOLD conf file tick box is activated e Hit Save then OK to overwrite the existing gold conf file GOLD will then start running interactively e The GOLD output window is a tabbed view that allows you to inspect various files that are written while the docking proceeds Once the job is complete the message Finished Docking Ligand ligand mol2 will appear in the gold_ligand_m1 log tab 160 GOLD User Guide Protein Log File e Inspect the gold_protein log file by hitting the gold_protein log tab i
112. SU Eireren eee e eee ea ea iteactwectevvanniteaetueesenveuneteeeteeds 226 23 3 N Mber of Operations ecne vee eek va ot ws Seas on deve ee vs Eea araar Eeee 226 23 4 Number of islands savant settee tie teen eee ee eee des 226 23 90 INIGNE SIZ awe sea AAA aA A A AeA AA BANA AA ee 227 23 6 Operator Weights Migrate Mutate CroSSOVES cccccccsssssssseeeeeeeesaeeseeeeeees 227 Appendix E Utility Programs ccccccccssssssseeecceeeeaeeeeeeeeeeeeesaaaeeseeeeeeeessaaaaseeeeeeeesaaa 228 QA SIMA IMS errete aE E E E vee vate eed eve toeec a e pas ETa 228 24 2 rms SAMALWSIS sore ae wees oles weds ease e e e a ea Seadoo ca dene era a T 229 243 identity ligand Pyer ETEEN RR A TE 230 DAA check MOND Exe ereere riii ti eiii eii i i a Pee 230 PK is eames 10 0 E 8 EE ere E E E E E E E EES 231 24 5 1 gold_utils protonate 2 0 cceccccecccccecseseeeeeceeeessaueesseeeeeesessaaaaess 231 24 5 2 gold_utils print_rotaMer ccccccccccccccssssseseceeccecaeaeeseeeceeeesaeaaeess 231 24 5 3 GOIA_UtIIS CONVETE 0 cccccssssseeeccceeeeaeeeeseeeceeceessuaesseeeeeeeesaaaaeess 232 GOLD User Guide 24 5 4 gold_utils write_complexes CONF cccccccecccececeessseeeeeeeeeeeaaeeees 232 25 Appendix F The Torsion Angle Distribution File cccccccccssssssseeeeeeeeseaeeeeeeseeeeeeseas 233 25 1 Format of Torsion Angle Distribution File Header ccscsessecceeeceseeeseseeeees 233 25 2 Format of Torsion Angle Distrib
113. Setup window You can then highlight the atoms belonging to the subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area Only those atoms specifically included in the binding site definition will be considered during docking The binding site definition should therefore be large enough to contain any possible binding mode of the ligand and include all atoms or residues that might be involved in ligand binding Since this binding site definition might include atoms that lie outside the cavity i e on the surface of the protein you can use cavity detection to restrict the binding site definition to concave parts of the binding site surface see Cavity Detection Each atom in the defined binding site is tested for solvent accessibility this is a two step process First the solvent accessible surface of each atom in the defined binding site is calculated Potential donor and acceptor fitting points used for ligand placement are then generated for only those protein atoms that are accessible Second the potential fitting points are themselves tested for solvent accessibility and only those fitting points that are accessible are used Therefore for a protein atom to be recognised as a donor or acceptor it must be included in the binding site definition be solvent accessible and have at least one associated solvent accessible fitting point It is possible t
114. T a ae 6 OP Be a aL 0 25 1 23 4 5 6 7 8 9 10 0 38 1 23 4 5 6 F7 8 9 10 0 64 Lo 2003 4 8 6 FB 29 P20 0 90 18 2 3 Ep eS 6 To 9 T a 1 52 18 9 2 3 4 5 6 7 10 1 95 ToS 6 T OB 9 es Be A a 4 50 15 6 7 8 9 2 3 410 6 37 123 4 5 6 7 8 910 e itis recommended that the Allow early termination tick box is disabled in the GOLD front end when generating diverse solutions see Early Termination e Itis possible to generate links to the top ranked solution from each distinct cluster at a given RMSD cutoff see Identification of Different Binding Modes Clustering of Ligand Poses GOLD User Guide 10 Setting Constraints 10 1 Using the Constraint Editor GOLD User Guide Depending on what sort of contraint s are required they may be protein specific or applied to a protein ensemble Protein specific constraints are the following Distance constraint for use with individual ligands see Distance Constraints Substructure based distance constraint for use with multiple ligands that have a common substructure or functional group see Distance Constraints Hydrogen bond constraint for specifying a hydrogen bond between a particular ligand atom and a particular atom in the protein see Hydrogen Bond Constraints Protein hydrogen bond constraint for specifying that a particular protein atom should be hydrogen bonded to the ligand but without specifying to which ligand atom
115. The docking program can then be used to predict the binding mode of the ligand and a comparison made with the crystallographically observed position The crystallographically observed conformation of the docked N phosphonacetyl L aspartate ligand is stored in the ligand we extracted from the protein that was subsequently re loaded A 1ACM in the Molecule Explorer Compare this with the solution predicted by GOLD e Inthe figure below the crystallographically observed reference structure A 1ACM shown in green is compared with the top ranked solution predicted by GOLD shown coloured by element e Using this methodology GOLD has been validated against a large number of protein ligand complexes taken from the PDB Further details and the entire validation test set are available for download This ends the tutorial 156 GOLD User Guide 20 2 Tutorial 2 Handling of Metals in GOLD Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial2 toa directory to which you have write permissions The object of this tutorial is to investigate the binding mode of brinzolamide an inhibitor of carbonic anhydrase II PDB entry code 1A42 In this example the brinzolamide molecule is known to coordinate to a zinc atom within the ligand binding site of the protein This tutorial will illustrate the requirements for setting up and running a docking in which the protein binding site features a metal ion Additi
116. This tutorial will illustrate how to use GOLD to generate diverse solutions and show how this feature can be used to improve the outcome of docking methylparaben insulin back into its native protein Preparation of Input Files The original PDB file 3MTH pdb has been provided should you wish to set up the protein and ligand files yourself Protein and ligand files are also provided and have been set up in accordance with guidelines for the preparation of input files see Setting Up the Protein s and Setting Up Ligands respectively These files can be opened in Hermes and inspected You will be able to see the extent of the protein active site compared to the size of the ligand GOLD Configuration Files Two GOLD configuration files are provided in lt install_ dir gt GOLD Suite GOLD examples tutorial8 The settings in these files and how to run the files is covered in the sections that follow gold conf see Running gold conf and Viewing the Results diverse conf see Running a Diverse Solutions Docking and Viewing the Results Running gold conf and Viewing the Results 198 The gold conf configuration file contains settings for carrying out a standard docking i e without generating diverse solutions Output files have already been generated for this docking and are provided in the gold directory These results can be viewed GOLD User Guide directly by opening Hermes and reading in the gold conf via Load GOLD res
117. Torsion Angle Distribution Files Copy the default file torsion angle distributions file that is provided in the GOLD_DIR gold directory to the current directory After making your changes instruct GOLD to use the edited file see Enabling Use of Torsion Angle Distributions The format of entries in the file is quite strict incorrect editing of the file may cause GOLD to behave in unexpected ways or even to crash For further information refer to see Appendix F The Torsion Angle Distribution File Matching Torsion Angle Distributions at Run Time GOLD identifies each rotatable bond in the ligand and attempts to match it to a torsion angle distribution in the torsion angle distribution file This includes bonds that are identified by GOLD as flippable e g if torsions are switched on then ligand carboxylic acids O C OH will also use a torsion distribution Details of matched torsions are written to the gold_ lt ligand_ name gt _ml 1log file specifically Anitemised list of which torsions have been matched during ligand initialisation including the torsion name e g Rotatable bond 40 41 61 63 matches torsion ester Cr Os 3 CRA 1C SOR2 4 E38 Rotatable bond 65 64 63 61 matches torsion acid T1 C22 Orco Orco i CoS C 2H lt Ga3e 28 2 jie Rotatable bond 67 65 64 63 matches torsion acid T2 Ores C2 e OLCO R C88 6 52H IEG BBC Matched torsion angles are now identified in the rotatable ligand
118. UNS ccccccccssssseseeeeeeeesaeaeeess 131 15 3 Comparison of Docking Solutions ccccccccccccccceeeeeeeececeeecseeeeseeeeeeeessaeaaness 131 15 4 Identification of Different Binding Modes Clustering of Ligand Poses 132 15 5 Viewing Docked Solutions in Hermes cccccccscsssssseeecceeeecaaeeeeeeceeeessaaaaness 134 GOLD User Guide vii viii 16 17 18 19 20 21 22 23 24 15 6 Analysing Results in GoldMine ccccceesseeecccccecceeeeseecececessaaeeseeeeseessaaaanees 134 15 6 1 Overview of GoldMine oe eeeceesscceeeeeeeeeeeeesaaeeeeeeeeeteteeaaeeeeeees 134 15 6 2 Sending Docking Results to GoldMine cccccccccceeccseeeeeeeeeeeeeeees 135 Saving and Reusing Docking Settings ccccccccccccccaeseseeeceeeeeeeeeeseeeeeeeesseaeaseeseeess 137 16 1 Saving and Re using Program Settings in Configuration Files ccceeee 137 16 2 Using Configuration File Templates ccccccccscsssssseeecceeeceeaeseeeceeeessaaaaeess 137 16 3 Customising Scoring Function ParaMetelS cccccccsseeceeceseseeceeeeeesseeeeeees 138 16 4 Customising the Torsion Angle Distribution File ccccccssssseeeeeeeeeeeeaeeeees 138 Context Dependent Helpz anc nae te See A E Eee 139 RETERENCCS 8 AEA EE E E E E EE E A EE E EAE 140 Acknowledgements mereen a a a a e aaar aieia 142 Appendix A Tutorial cccccsssccceeccccsccesssseeeecccesaeasseeeeeecceesaaaseceeseccesa
119. Using Torsion Angle Distributions GOLD will attempt to match amide torsions against the torsion angles distributions file If an amide torsion matches this will override the Flip amide bonds flag setting Note Data in the CSD show that both cis and trans conformations occur in ureas it is therefore recommended that amide flipping be turned on in order to sample R N C O N torsions of 0 degrees when docking ureas 8 3 Flipping Pyramidal Nitrogens Click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Flip pyramidal N check box to allow pyramidal i e non planar sp3 nitrogens to invert during docking otherwise they will be held fixed at the input geometry Given a non planar group RR R N or tetrahedrally surrounded RR R NH the Flip pyramidal N switch enables flipping of the local stereochemistry around the nitrogen the energy barrier for this umbrella like change of geometry around the nitrogen is low Flipping only changes the stereochemistry around RR R N and RR R NH nitrogens It does not affect other chiral centers 8 4 Intramolecular Hydrogen Bonds 80 Click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Detect Internal H bonds check box to allow intramolecular hydrogen bonds in the ligand to be formed during docking GOLD User Guide e Use this with care as it can make
120. a Point GOLD User Guide This binding site definition might therefore include atoms that lie outside the cavity i e on the surface of the protein You can use a cavity detection algorithm Hendlich Rippmann and Barnickel LIGSITE Automatic and efficient detection of potential small molecule binding sites in proteins Merck technical report 1997 to restrict the region of interest to concave parts of the binding site surface To enable cavity detection switch on the check box labelled Detect cavity restrict atom selection to solvent accessible surface This option is available by clicking on Define Binding Site from the list of Global Options given on the left of the GOLD Setup window After docking the atoms included in the binding site definition are listed in the Cavity atoms section of the gold_protein log file The cavity atom selection is also saved as a protein atom subset within Hermes You can highlight the atoms belonging to any subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area It is possible to generate contour acnt files of the cavity used by GOLD by editing WRITE CNT FILES 0 to WRITE CNT FILES 3 inthe gold params file see Altering GOLD Parameters the gold params File The acnt files produced can be read into Hermes following the docking via Display Contour Surfaces Further details on how to read acnt files into Hermes are provid
121. add a new ring template not already present in the CSD derived library or if you wish to override the CSD conformations for an existing template First you must create a ring template library file in the directory lt GOLD DIR gt gold user_ ring conformations template library mol2 This mol2 file should contain all user defined ring types Rings in the ligand are matched against the rings in this template file The atom types in the template_library mol2 file must therefore match the ligand atom types exactly The molecule identifiers inthe template library mol2 file must start at 0010001 The identifier must be incremented by 1 for each successive ring in the file For example the following template library mol2 file contains two templates a triazole and a cyclohexane ring GOLD User Guide lt TRIPOS gt MOLECULE 0010001 5 5 aN D 0 SMALL NO_ CHARGES KEKK triazole lt TRIPOS gt ATOM 1 N pl3 0 0098 7 5008 14 4731 N pl3 1 RES1 0 0000 2 C22 0 6346 8 7211 14 5033 cC 2 1 RES1 0 0000 3 N 2 1 7255 8 6751 13 7686 N 2 1 RES1 0 0000 4 N 2 1 8102 7 3942 13 2152 N 2 1 RES1 0 0000 5 6 2 0 7618 6 7331 13 6432 C 2 1 RES1 0 0000 lt TRIPOS gt BOND 1 3 2 2 2 4 3 a 3 5 4 2 4 1 2 1 5 5 aR at lt TRIPOS gt MOLECULE 0010002 6 6 1 0 0 SMALL NO_CHARGES KKKK cyclohexane lt TRIPOS gt ATOM 1 2 5 1537 5 6169 11 3626 C 2 1 RES1 0 0000 2 C 3 4 2377 4 7380 11 9725 C3 1 RESL 0 0000 3 3 4 7266 4 0400 12 9517 C 3 1 RES1 0 0000 4C 3 6
122. al Xmax e Inthe GOLD implementation of ChemScore the block function is sometimes convoluted with a Gaussian function Tee a e u o du il Se See mene 0 2 Be ideal max T g u o du g u O peio e The effect is to smooth the function e g 62 GOLD User Guide Xideal Xmax 7 4 3 Hydrogen Bond Terms e The hydrogen bond term is computed as a sum over all possible donor acceptor pairs such that one atom belongs to the protein and the other to the ligand e Each term in the summation is the product of three Gaussian smoothed block functions see Block Functions in ChemScore The purpose of the block functions is to reduce the contribution of a hydrogen bond according to how much its geometry deviates from a ideal H A distance b ideal D H A angle and c ideal directionality with respect to the acceptor atom The maximum contribution of a given donor acceptor pair to the summation is 1 this will occur if the pair form a hydrogen bond of ideal geometry AG LB Ar Ar Ar_ 0 B Aa A _ AQ_ 0 B AB AB AB 0g hbond alldonor acceptor pairs e The tables below describe the various parameters in this equation their meanings and what they are called in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File D H A distance parameters D Donor A Acceptor Term Meaning Name in ChemScore File Default Value r The ideal hydrogen accep
123. alues for other rotamers can be viewed in the dials by clicking on each Rotamer line in turn The settings on the dials describe the following green the observed torsion in the protein red the defined rotamer Chi value that will be used during docking blue wedge the tolerance allowed Delta for the defined rotamer e There are a number of other options for setting rotamers other than the library settings Rigid this fixes a particular side chain at its input conformation i e makes it non flexible during docking Free this allows a side chain to rotate freely during docking i e the defined rotatable torsion will be permitted to vary over the range 180 to 180 Crystal this setting will define a rotamer in which all rotatable torsions in the side chain will be allowed to vary over the range delta chi to delta chi where chi values are taken from the protein input file From dials this allows rotamers to be specified directly Start by setting each chi value click on the dial and while holding down the mouse button move the red indicator line to the required position The corresponding torsion will rotate within the Hermes visualiser to show the current value Alternatively type the required chi value into the entry box directly under the dial When all chi values are as required press the From dials button to accept the rotamer definition e We will use the Library settings for this docking so if you ha
124. ances Identify any additional hydrogen bonding interactions between the benzimidazole substituents and the target subsites within the protein This ends the tutorial GOLD User Guide 171 20 4 Tutorial 4 Use of Substructure Based Distance Constraints Introduction e First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial4 toa directory to which you have write permissions e The object of this tutorial is to assess the binding of a small number of structurally related ligands with the carbonic anhydrase II PDB entry code 1cil In the ETS inhibitor a terminal sulphonamide nitrogen atom is observed to coordinate to a zinc atom within the protein binding site e This tutorial will illustrate how GOLD can be used to screen a number of compounds in order to identify ligands with potential activity The use of constraints in order to bias solutions towards the observed binding mode of the inhibitor will also be demonstrated as well as the use of automatic speed settings Input Files e Open Hermes and read in the file protein mol2 from the folder to which you copied the tutorial4 files The original protein PDB file 1C IL pdb has also been provided should you wish to set up the protein for yourself e Carbonic anhydrase II 1cil protein mo l2 has already been set up in accordance with the guidelines for the preparation of protein input files see Protonation and Tautomeric States e Upon inspection of the
125. and complex its intrinsic binding affinity needs to outweigh the loss of rigid body entropy on binding Where waters are specified in the gold conf an additional parameter S bar is added to the fitness score calculation Fitness S hb_ext 1 3750 S vdw_ext S hb_int 1 0000 S int S bar S bar is a barrier penalty term associated with non displacement of water 3 5 2 10 Specifying Waters GOLD allows you to retain specific water molecules that are important to ligand binding i e you can specify whether a particular water should be present or absent in the protein Furthermore for waters which are retained GOLD can automatically determine whether a water should be bound or displaced by the ligand during docking i e by toggling it on and off during the run The orientation of the water hydrogen atoms can also be optimised by GOLD during docking In addition the location of each water molecule can be allowed to translate within a radius of 2 A Click on Configure Waters from the list of Global Options given on the left of the GOLD Setup window Waters must be specified in separate files i e one water per mol2 file To specify the water files select the Add button at the bottom of the Configure Waters window Use the file browser to locate the water files select one or multiple files then hit Open to add them to the Configure Waters dialogue If the protein file contains all waters i e active and non active w
126. and seems to offer lots of scope for users to commit errors For that reason we recommend that the PDB format is not used for ligands Specifying the Ligand File s Any number of ligands can be specified either by selecting several individual files or by selecting a single file containing several ligands i e a mu ti MOL2 or SD file GOLD will dock each in turn Acceptable ligand file formats are MOL2 i e Tripos format MOL i e MDL SD format see Ligand File Formats Click on Select Ligands from the list of Global Options given on the left of the GOLD Setup window 41 3 GOLD Setup See Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Ligand File GA Runs First Ligand Last Ligand 1 ligand mol2 Proteins Define Binding Site Select Ligands Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing J Show full file paths Add Delete Reference ligand SD Help R Run GOLD Run GOLD In The Background Finish Cancel e To specify a ligand file click on the Add button and use the file selection window to choose the ligand data file s e Specify the number of times each ligand is to be docked by entering a value in the GA runs box see Number of Dockings e When using a single file containing several ligand
127. arch problem is not large and the same settings can be used throughout Hit Output Options and either browse to or type the name of an appropriate sub directory in the Output directory box e g waters_toggle Hit the Run GOLD button at the bottom of the GOLD front end In the Finish GOLD Configuration window you will be prompted that the GOLD configuration has been updated and needs to be saved GOLD User Guide 181 We have not modified the protein mol2 file so we do not need to save this file thus ensure the tick box adjacent to protein mol2 is deactivated Change the configuration file name to e g waters_toggle conf then hit Save to start the GOLD run This will start the GOLD job interactively As the job progresses output will be displayed in the Run GOLD window Once the job is complete the message Finished Docking Ligand ligand mol2 will appear in the gold_ligand_m1 log tabbed view of the Run GOLD window Hit Close in the Run GOLD window to close it Running GOLD Dockings All waters turned on Return to the GOLD front end and click on Configure Waters to bring up the water setup window Change the toggle state of each water molecule to on Go to Output Options and change the output sub directory name e g to waters_on Hit the Run GOLD button In the Finish GOLD Configuration window as before edit the name of the GOLD configuration file in the GOLD conf file text box to e g waters_on conf Hit Save Once the GOLD r
128. ate large ligands In such cases it is possible to apply a softer Split Van der Waals Potential for certain selected residues Two alternative soft Split Potential forms are parameterised in the gold params file Potential 1 EXTERNA POTENTIA 1 4 2 Potential 2 4 8 EXTERNAL POTENTIAL 2 4 8 2 i The first term of each form describes long range interactions the second term describes short range interactions The point of change over is at the 4 8 potential minimum and the second term is set such that both terms take the same value at this point The function therefore remains continuous and the minimum point is the same as with the default 4 8 potential Soft potentials are protein specific so to apply an alternative soft potential to specific residues you must first activate the protein tab adjacent to the Global Options tab e g Protein 1fax coagulation factor in the example below then click on Soft Potentials from the list of available options given on the left of the GOLD Setup window gt GOLD Setup oix Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial flexible conf Load Save Options Protein 1fax coagulation Factor Protonation amp Tautomers Waters Delete Ligands Residues alternative potential 1 Flexible Sidechains Soft Potentials Metals Constraints Covalent Interaction Motif
129. aters the active waters must be extracted and the non active waters deleted in the following way Click on the protein name tab adjacent to the Global Options one in the example below this is the 7ACM tab then select the Extract Delete Waters option 3 GOLD Setup BBE Conf file ee load Save Global Options 14CM Protonation amp Tautomers Extract Delete Waters Waters Delete Ligands Flexible Sidechains Soft Potentials Metals Constraints Covalent Interaction Motif Extract Waters For Docking Delete Remaining Waters Waters that are important for ligand binding can be added to the list of active waters used in docking by checking them in the water list above and clicking the Extract Waters For Docking button Once you have extracted the waters you wish to make active press the Delete Remaining Waters button to remove all waters from the protein Help R Run GOLD Run GOLD In The Background Finish Cancel Select the waters you wish to keep either by selecting them in Hermes 3D view or by activating their corresponding tick box Selected waters can be GOLD User Guide unselected by deactivating their tickbox or by deselecting them in Hermes 3D view Hit the Extract Waters for Docking button This will write the waters to individual files in the working directory The files will have names of the type 1ACM_HOH15 mol2 Once you ha
130. ations The torsion angles around rigid bonds such as amide linkages double bonds and certain bonds to trigonal nitrogens will normally be fixed at their starting values However you can use the Ligand Flexibility options to enable some of these features to vary see Ligand Flexibility GOLD will not alter stereochemistry If you are unsure about the stereochemistry of the ligand you must generate all alternatives and dock each separately It is meaningful to make comparisons between fitness scores for dockings of different stereoisomers Ligand File Formats Acceptable ligand file formats are MOL2 i e Tripos format MOL i e MDL SD format and PDB although we do not recommend the use of pdb format Files in MOL format may also have the extension mdl or sdf Only MOL2 may be used if you wish to set ligand atom types manually see Automatically Setting Atom and Bond Types An extension to the PDB file format is required if it is used for storing the ligand structure Specifically a bond specified twice in a single CONECT record is assumed to be a double bond and a bond specified three times in a single CONECT record is assumed to be a triple bond For example the following CONECT records both specify a double bond between atoms with serial numbers 25 and 26 CONECT 25 20 26 30 26 CONECT 26 25 27 52 25 This mechanism for specifying bond orders is forced by the lack of a bond order field in the standard PDB format
131. atom ID GOLD User Guide GOLD Setup 1905 Note that you can toggle the ligand protein on and off via the Molecule Explorer to help make the selection of both covalent constraint atoms easier BBs Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Waters Help R Protonation amp Tautomers Delete Ligands Flexible Sidechains Protein link atom 1905 NZ LYS258 Soft Potentials Metals i ag iil Ligand link mode Atom Substructure Substructure HBond Protein HBond Ligand link atom 25 Nz RES1 Covalent Interaction Motif Substructure file RLY E E EFF_ Hs IV Define covalent docking VV Use topology matching to check test equivalent atoms You can define the covalent link atoms by right clicking in the viewer or by typing in the edit box The ligand link atom can be defined either by a single atom in the ligand or by an atom in a substructure that can be matched against multiple ligands If using a substructure you must enter the substructure file Run GOLD Run GOLD In The Background Finish Cancel Once you have finished defining the covalent atoms click on the Global Options tab to return to the general docking setup Click on Output Options and specify a directory to which you have write permission This will be where the output
132. ays The following commands are recognised in PLP parameter files For the default values used in PLP and CHEMPLP see the plp params and chemplp params parameter files respectively Parameters used for PLP and CHEMPLP plp params and chemplp params PLP_COEFFICIENT Weight of PLP contributions Wp PLP_LIGAND_CLASH_COEFFICIENT Weight of ligand clash potential Wyig ciash PLP_LIGAND_TORSION_COEFFICIENT Weight of ligand torsion potential Wyig tors PLP_PROTEIN_ENERGY_COEFFICIENT Weight of ChemScore protein potential Wpror PLP_CONSTRAINT_COEFFICIENT Weight of constraint contributions Wons PLP_GRID_SPACING Grid spacing used for PLP map PLP_HBOND_METAL_FUNCTION Additional hydrogen and metal bonding contributions Use CHEMSCORE to activate ChemScore hydrogen and metal bonding contributions If NONE is specified only the PLP contributions will be considered see the plp params file PLP_WATER_BARRIER Penalty value added for each explicit water molecule activated by the search algorithm positive value HBOND_A HBOND_B HBOND_C HBOND_D HBOND_E lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt CHEMSCORE NONE gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt GOLD User Guide Parameters used for PLP and CHEMPLP plp params and chemplp
133. best solution from the flexible run has a much higher GoldScore value 76 8235 than was obtained from the rigid run The movements of the flexible side chain GIn192 can be seen more effectively if the representation style of the GlIn192 residue is changed Click on Selection then Define Complex Selection Activate the By residue radio button then hit the Specify Individual button at which point you can select GLN192 from the list of residues Click Add Close Save then Close to complete the selection The newly defined subset can be selected by picking it from the Atom selections pull down menu in Hermes The display style can be changed after the residue is selected by right clicking and selecting Styles from the pull down menu Choosing Side Chain Rotamers 192 Two decisions must be made when using the flexible side chain facility a which side chains are made flexible b how flexible is each side chain made It is important to recognise that the more flexibility is introduced the larger the search space becomes Particularly with high throughput runs when relatively little time can be allowed per ligand this may seriously decrease the chance of finding the global minimum A sensible strategy is therefore to make a side chain flexible only if you have some a priori reason to suppose that it will move as we have from X ray structures in the tutorial example On the other hand we probably allowed GlIn192 more movement than neces
134. bility SSIS Fitness amp Search Options User specified list GA Settings _ZwZw__yOOOOONn 7 Output Options GoldMine Parallel GOLD i E dky Centre of sphere region z9 5i oss poll Radius of sphere Angstrom B O egion Atom Typing Define sphere Score contribution per atom Found in region 1 0 J Never dock a ligand when a constraint is physically impossible ibe Det OSS Region 2 94 5 1 0 69 3 1 0 hydrophobic_atoms Edit Sphere 21x Create or Edit Sphere Name Jconstraintt Radius 3 Position Sphere at Centroid of protein subset Add subset Centroid of atoms selected in 3D viewer coordinates x 2 937 y fs 103 z 0 686 IV Centroid visible Cancel 4 e Enter a name and the radius of the sphere distances are in A e The sphere must then positioned within the binding site this can be doneina number of different way The sphere can be positioned on the centroid of an existing subset of protein atoms Select Centroid of protein subset and select the protein subset from the drop down list To create a new subset of protein atoms click on the Add Subset button Instructions on defining protein subsets can be found in the Hermes user guide The sphere can be positioned on the centroid of selected atoms Select Centroid of atoms selected in the 3D viewer then within Hermes click on one or more protein atoms in order to define a centroid Alternatively select
135. bstructure Based Covalent Links It is possible to apply a covalent link to multiple ligands which have a common functional group During docking the link will be applied to any ligands which contain a specified substructure matching is performed on the basis of the atom types and 2D connectivity Note the substructure must be a sub graph rather than a complete molecule To use a substructure based covalent link first create a file containing the substructure in MOL2 format e g substructure mo1l2 It is recommended that you set atom types manually see Manually Setting Atom and Bond Types since an incomplete fragment can cause problems with automatic atom typing The actual conformation of the group in this file is not important as only the atom types and 2D connectivity will be used Covalent constraints are specific to the protein thus click on the protein tab e g Protein 1ase aminotransferase in the example below select Covalent from the list of available options given on the left of the GOLD Setup window and enable the Define covalent docking check box Select Substructure as the ligand link mode GOLD User Guide 3 GOLD Setup mE Ea Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Protonation amp Tautomers Waters IV Define covalent docking Delete Ligands Flexible Sidechains Protein link
136. c iron parameters in the context of docking to heme containing proteins and demonstrated improved performance It is now possible in GOLD to optionally use these parameters The parameters are derived from contact statistics obtained from the CSD and PDB databases Parameters were derived for both GoldScore and ChemScore These parameters can be used by choosing the appropriate scoring function params file from those that have been supplied with the GOLD installation The scoring function params files that are available are goldscore p450_csd params goldscore p450_pdb params chemscore p450_csd params 75 76 chemscore p450 pdb params e The files are located within the SGOLD_DIR gold directory The graphic below shows the iron parameters for GoldScore derived from the CSD as displayed in the goldscore p450_csd params file WY HEIE SCOURS FORT W Ber RLrton Eet al E DETAL COMED MATION CS TAL caver TAL CUORI TAL CARI ra E n ala ale THE FRE EEE a De be Da Da Da Da be ba bw 00 00 00 00 00 00 00 00 00 Prove ne 5 7 6 7 HS 2 6 5 5 0 3 9 _ e Fe re re Fe re Fe Fe re Fe e005 5S pps 6 44 e To employ one of the files click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select GoldScore or ChemScore from the Scoring Function drop down menu Then either enter the path and filenam
137. can be changed by altering the parameter CLASH RADIUS _HBOND in the ChemScore file see Altering ChemScore Fitness Function Parameters the ChemScore File Any metal coordination contact shorter than rmeta A contributes a clash term of _ 20 0x r metal clash metal G metal metal The value of rmeta default 1 3 A can be changed by altering the parameter CLASH RADIUS METAL in the ChemScore file see Altering ChemScore Fitness Function Parameters the ChemScore File All other ligand protein interatomic contacts contribute clash terms of the following form clash 40 Waari clash other clash lciash Varies with contact type for contacts to protein sulphur atoms it is set to 3 35A for all other contacts it is set to 3 10A These settings correspond to the parameters CLASH RADIUS SULPHUR and CLASH RADIUS GENERAL in the ChemScore file see Altering ChemScore Fitness Function Parameters the ChemScore File Internal ligand strain is accommodated by clash terms in combination with torsional strain terms of the form 67 DA cos n All rotatable bonds internal e Bonds are deemed to be rotatable if they are single and acyclic and involve pairs of atoms with hybridisation states sp3 sp3 sp3 sp2 or sp2 sp2 e The parameters A n and in the above equation are set in the ChemScore file see Altering ChemScore Fitness Function Parameters the ChemScore File The relevan
138. cantly favoured over the other in a way you do not intend When using improper torsions These distort the protein structure in artifactual ways and will often introduce an artifactual protein protein clash 4 2 Large Backbone Movements It is not possible for GOLD to make large backbone movements This sort of problem can be dealt with by performing an ensemble docking see Ensemble Docking Small backbone movements in the vicinity of a flexible side chain may be allowed by including the improper torsion angle CA N C CA in a rotamer_lib command block see Allowing a Localised Backbone Movement Another option you can try is to apply a Localised Soft Potential to one or more residues in the loop see Allowing For Localised Movements Docking With Soft Potentials 4 3 Ensemble Docking 4 3 1 Introduction Sequential docking of individual ligands into a protein is computationally time consuming Ensemble docking aims to address the issue of protein flexibility by adding multiple protein structures into a single GA run The ultimate aim is to obtain higher enrichments in virtual screening experiments Multiple protein conformations can be searched concurrently when docking an ensemble thus saving valuable time compared to a sequential docking approach Starting from a superimposed set of protein structures GOLD evolves a separate population of individuals representing ligand conformations for each protein structure part of the ens
139. ccccssesseeecceececsaeeesseeeceeeesseaeeeeeeeeeeesauaaaeeseeeeeeeaaa 33 4 3 1 NIL ROCU CUI ON eresie seet EE EE LAE KEL AERES 33 4 3 2 Setting up Proteins for Ensemble Docking ccccccceeeeeeeseeeeeeeees 33 4 3 3 Setting up an Ensemble Docking ccccccccecseessseeeeeceeeesaaaeseeeeeees 34 4 3 4 Interpreting Ensemble Docking Output ccccccssssssseeeeeeeeseaeeeees 36 4 3 5 Caveats of Docking into Ensembles cccccccssssseseeeeceeeeeaaaeseseeeees 37 4 4 Allowing For Localised Movements Docking With Soft Potentials 37 S tting UP Ligands veces cc cececvesceecceecasescpeeebesctatetusobadcGee Tera ebad chebatababeebetehabcdnbedebeded bance 40 5 1 ESSCN tial E PS E 40 5 2 Ligand Hydrogen Atoms lonisation States and Tautomeric States 0 40 5 3 Ligand Geometry Conformation and StereochemMistry cccccsssseseeeeeeeeeeeaes 41 5 4 Ligand File ForMats 22024 can acten RAwA ee nn nnn AA A a 41 5 5 Specifying the Ligand File s ccssssssssssssssssessssseessessesessessssssesseesssessseeeaes 41 5 6 Receiving Ligands From GoldMine ccccsssesseeccccceceeaesseeecceeeesaeaeeeseeeeeeeeeaas 43 5 7 Setting Up Covalently Bound Ligands ccccsssssesseecceceeeaseseeeeceeessaaaaseeseeees 44 5 7 1 Method Used for Docking Covalently Bound Ligand6s 000088 45 5 7 2 Setting Up a Single Covalent Link ccccccccccsssseeeeceeeeeeaeaeseee
140. ce alignments which give a pair wise matching of one residue to another which can then be used for overlay Use 1t9s as the reference chain and choose to overlay 1TBF only Click Next to proceed to the Protein setup dialogue GOLD Setup EF wizard step 2 Protein setup At this point you have the chance to edit your protein structure if required e g add hydrogens delete waters Global Options 1TBF 1t9s 1xp0 2chm Wizard steps To edit the protein s use the options available on the protein tabs to L Siet apai 1 Add Hydrogens 2 Protein setup 3 Define the binding site 2 Configure active waters and delete unnecessary waters 4 Configuration template 5 Select ligands 3 Delete ligands 6 Choose a fitness function 7 GA search options 8 Finish Help R lt Back Next gt Cancel Wizard Since the proteins are now set up hit Next again to proceed to the Define the binding site dialogue Defining the Binding Site 212 It is necessary to specify the approximate centre and extent of the protein binding site Since binding site definition for an ensemble must be a position suitable for all proteins it is not possible to define the binding site from an atom or a list of atoms or residues It is only possible to define the binding site from a point in space or from a ligand Open the 1x0z ligand in Hermes by using File then Open and then selecting the 1x0z mo12 file This makes this ligand available for binding sit
141. chines smart_rms calculates the rms difference between two conformations of the same structure while taking account of symmetry effects such as the flipping of a phenyl ring by 180 degrees Using a graph isomorphism algorithm an rms score is calculated for each way of mapping the molecule onto itself smart_rms can be invoked from the command line The following platform dependent commands should be used Linux platforms lt install_dir gt GOLD_Suite bin smart_rms hv conformation_1 conformation_2 Windows platforms at the Windows command prompt lt install_dir gt GOLD gold d_win32 bin smartrms_win32 exe hv conformation_1 conformation_2 where lt install dir gt is the GOLD installation directory If specifying the full path the command will need to be in inverted commas e g C Program Files CCDC GOLD_Suite GOLD gold d_win32 bin smartrms_win32 exe hv conformation_1 conformation_2 The flags are h use heavy atoms only the calculation easily becomes intractable if Hs are included v verbose output conformation_1 and conformation_2 are MOL2 files containing the two conformations GOLD User Guide 24 2 rms_ GOLD User Guide analysis Located in C Program Files x86 CCDC goldsuite 5 3 GOLD gold d_win32 bin on Windows machines rms_analysis calculates an rms difference matrix for a set of structures as MOL2 files and performs hierarchical cluster analysis A graph isomorp
142. cking a wizard is available which will guide you through the essential configuration steps Select GOLD from the top level menu in the Hermes visualiser then Wizard from the resulting menu Alternatively the wizard can be opened at any stage from the GOLD Setup window by clicking on Global Options on the left of the window then clicking on the Wizard button gt GOLD Setup Me Ea Wizard step 1 Select one or more proteins Either choose a protein already loaded in the visualiser or load a new file Options Wizard steps N 5 S A 1 Select a protein Select proteins to use Load Protein Superimpose Proteins 2 Protein setup 3 Define the binding site 4 Configuration template 5 Select ligands 6 Choose a fitness Function 7 GA search options 8 Finish Protein score offset ensemble docking only lt Back Next gt Cancel Wizard e The appearance of the wizard will vary depending on whether a protein file has been read in or not General docking settings are available from within the Global Options tab while protein specific options only become available after a protein has been loaded into Hermes either via File Open or via a gold conf These protein specific options can be found under an additional tab found next to the Global Options tab The text on this tab is taken from the HEADER record ina PDB file or the lt TRIPOS gt MOLECULE record ina mol file e The number of tabs wi
143. cov considering flexible side chains fchem prot and explicit water molecules as well as handling constraints eons Parameters for both fitness functions can be altered by changing the files plp params and chemplp params for PLP and CHEMPLP respectively see Altering PLP Fitness Function parameters CHEMPLP parameters are used by default GOLD User Guide as they show on average an improved performance in pose prediction and virtual screening applications fitnessprp Weip forr Wiig clash ig clash Whig tors Sisters fehem cov Wbprot fchem prot Weons Feons fitn ESS cHEMPLP fitn SSpip fchem hb chem cho ehem met 7 2 2 PLP Interaction Types e The Piecewise Linear Potential PLP models the attraction as well as repulsion of protein and ligand heavy atoms In Figure 1a the partially attractive potential using 6 parameters is presented Parameters A to D and in Figure 1b the purely repulsive potential using parameters A to D are presented e All protein and ligand heavy atoms are typed as donor acceptor donor acceptor or nonpolar Additionally metal ions in the protein s binding site are assigned the metal type e Depending on protein and ligand atom type the appropriate potential from Table 1 is selected Each potential H bond metal buried and nonpolar is defined by a specific setting of parameters A to F The same accounts for the repulsive potential in which case parameters A to D are s
144. dbigpe_full_entry Fitness amp Search Options pdb1gpj_full_entry GA Settings Output Options pdb3ick_a GoldMine Parallel GOLD Constraints Atom Typing T List all loaded files not just proteins Protein score offset ensemble docking only negative numbers favour a model positive numbers disfavour a model Score Offset pdbigpc_full_entry 0 0 T jo pdb1gpd_full_en pdbigpe_full_en pdb1qpj_full_entry 0 F Help R Run GOLD Run GOLD In The Background Finish Cancel e Proteins must be set up in the usual way and superimposed before they are using in an ensemble docking see Setting up Proteins for Ensemble Docking e From within the Protein window it is possible to apply a Protein score offset This user defined value will be subtracted from the overall fitness score if a ligand is docked into this protein structure Both negative and positive values can be used negative values favour the selection of a protein conformation positive ones disfavour thus using the protein score offset it is possible to bias which protein is selected There are no limits for these values If using this feature these scores are reportedas DE Protein inthe GOLD log files e When docking an ensemble the binding site definition must describe the binding sites of all loaded and superimposed proteins The binding site must therefore be defined using a method that isn t protein specific i
145. ding Docking Results to GoldMine e To receive ligands from GoldMine first it is necessary to create or load a valid GOLD configuration file Since it will often be the case that you will be using a tried and tested docking protocol to re dock selected ligands reading in an existing file is probably the usual case e Click on Select Ligands from the list of Global Options given on the left of the GOLD Setup window and delete any ligand file specifications made there e Click on GoldMine from the list of Global Options given on the left of the GOLD Setup window Within this pane it is possible to set GOLD up so that it either receives or sends docking poses from GoldMine e To receive ligands enable the Get ligands from GoldMine check box An appropriate machine Hostname and a Port number should be provided It is also possible to specify the Number of GA runs that will be carried out on each ligand The default is 10 3 GOLD Setup Jot x Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase r IM Get ligands from Goldmine Wizard Templates Proteins Hostname asphalt Define Binding Site Select Ligands Port aves Waters Ligand Flexibility Number of GA runs 10 Fitness amp Search Options GA Settings Output Options Goldmine J Send ligands to GoldMine Parallel GOLD E Constraints Hostname Jasphalt Atom Tyypin hiik Port aves
146. display can be controlled by using the settings under the Display tab Further options are available on right clicking v File Edit Selection Display View Calculate Descri Highlighting Depth Cueing Stereo Graphics Objects v Picking Mode Pick Atoms Clear Measurements dy 9 Atom selections ov Molecule Explorer 8 x Display Movable Descriptors n o E Z p 6 5 e 2 2 All Entries 4 vw Mw yw M hormone MMM 4 Bee Lal Styles Colours gt Labels Select Select Only Deselect Contact Management Auto Select is x Delete Define H Bond Center 3D view tg Center amp Zoom 3D view f Protein H Bonds Short Contacts 1 hormone methylparaben insulin GOLD User Guide 199 All docked solutions can be viewed by returning to the Docking Solutions window and pressing the Shift keyboard key whilst using the mouse to select first the top then the bottom solution You will notice that all the solutions are very similar None of the solutions replicate the original binding mode The ligand OH group is H bonding to the same CYS6 carbonyl group however the size of the protein active site means it is possible for the ligand to occupy an alternative area of the cavity than in the original crystal structure Also the water molecule known to mediate the protein ligand interaction is not being used by the pose above The RMSD of the top
147. dow e Switch on the button labelled One or more ligands A list of those ligands currently loaded in the Hermes visualiser will be shown Loaded ligands might typically include ligands in a known binding mode or the co crystallised ligand Select the reference ligand s you wish to use from this list Multiple ligands can be selected by left clicking whilst holding down the shift key gt GOLD Setup me Ez Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Atom select an atom in the visualiser or enter an atom index Proteins Define Binding Site zn ZN262 Yjew Select Ligands Waters Point select atoms to define a centroid or edit XYZ Ligand Flexibility Fitness amp Search Options GA Settings K 6 4223 y 0 8680 Z 15 2360 Views Reset Output Options GoldMine Parallel GOLD One or more ligands Constraints s Atom Typing 4 reference ligand from 14C List of atoms or residues Filename l rY View Select all atoms within 5 A J Generate a cavity atoms file From the selection Refine Selection IV Detect cavity restrict atom selection to solvent accessible surface J Force all H bond donors acceptors to be treated as solvent accessible Add Definition as a Selection Help R Run GOLD Run GOLD In The Background Finish Cancel e By defau
148. drogen Bond Correction ccccccccccccsssssseeeeeeeeseaaaeees 71 7 5 5 Covalent Docking and Docking with Constraints cccccssssssseeeees 72 7 5 6 Targeted Scoring Functions cccccccccccccssssseeeceeeeeeaeaeseeeeeeeesaaaaaess 72 7 5 7 Performance of the ASP fitness function ceccccccccccceeeeeeeeeeeeeeeees 72 7 5 8 Altering ASP Fitness Function Parameters the asp params File 72 7 6 User Defined Scoring Function cccccccccsssssseeeecceeeeaaaeeseeeeeeeesaaaaaeeeseeeeeesaaa 73 7 7 Altering GOLD Parameters the gold params Fil ccccccccccccesessseeeeeeeeeeeaes 74 7 8 Targeted Scoring FUNCIONS orrera esse ssesvecesd ert sonaa EEEE EEEE EEEE EEEE 75 7 8 1 Kinase Scoring FUN CtION 2e 2 ece eect et et ete a at eee 75 7 8 2 Heme Scoring FUNCTION cceccccceseeececeeeeceeeeseecseeeseeeaeeeseesaeeseeees 75 8 Ligatid Flexibility AE E 77 8 1 RING CONFONIMATIONS veces cecsseessees cece tect eeeesees seas eucdevedesdeeasteacdevedestseasteaduvetestieaieee 77 8 1 1 Flipping Ring COPNELS isser a aea ea Eia 77 8 1 2 Using CSD Ring Conformation Templates ccccccsssesseeeeeeeeeeeeeeees 77 8 1 3 The CSD Ring Conformation Library and Matching Templates at Run HM ieee ere ree rerreer reer n Ecce a a e a gE 77 8 1 4 User Defined Ring Conformations cccsccccccccecessesssseeseeeeeeeeaeeess 78 8 2 Flipping Amide Bonds cccccccssssssssecceececcaaeeeseeecceeessaaasseeeee
149. e Hit Next to proceed to the Finish window Specifying a Directory for GOLD Output 150 We are now finished our docking setup and represented with a Run GOLD button with which we can start the docking If we were to click on Run GOLD now the output would be written to the directory the 1ACM pdb file is stored in It is generally preferable to write output to a separate directory This option is available as part of the advanced options so rather that clicking on the Run GOLD button click on the Advanced button on the bottom right of the interface This takes us to the standard GOLD interface Select Output Options under Global Options This page is separated into three tabbed views File Format Options Information in File Selecting Solutions all of GOLD User Guide which allow control of which files are output and what information is written to the files GOLD Setup a Conf file Load Save Global Options 1ACM Wizard File Format Options Information in File Selecting Solutions Templates Proteins Output file format Same as input SD file C Mol2 Define Binding Site Select Ligands Output directory Configure Waters C Create output sub directories for each ligand Ligand Flexibility Fitness amp Search Options 7 Save ligand rank rnk files GA Settings a Output Options v Save ligand log files GoldMine v Save initialised ligand files Parallel GOLD Concniaie Save solutions to one fil
150. e Constraint contribution to PLP value See see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Protein Protein Clashes see Water Molecules see Internal Energy Offset see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP e Certain docking score terms are the product of a term dependent on the magnitude of a particular physical contribution e g hydrogen bonding and a scale factor determined e g by a regression coefficient e The docking score term descriptors included in the output file can therefore consist of weighted terms non weighted terms or both see Controlling the Information Written to Ligand Solution Files e Weighted terms will be indicated as such in the tag name e g Gold Chemscore Hbond Weighted GOLD User Guide 225 23 Appendix D Genetic Algorithm Parameter Definitions 23 1 Population Size The genetic algorithm maintains a set of possible solutions to the problem Each possible solution is known as a chromosome and the set of solutions is termed a population The variable Population Size or popsize is the number of chromosomes in the population If n_islands is greater than one i e the genetic algorithm is split over two or more islands popsize is the population on each island Changes to genetic algorithm parameters should be made with care see Controlling
151. e Atom Typing _ Use alternative bestranking Ist filename Create links for different binding modes based on RMSD clustering Help h Run GOLD Run GOLD In The Background Finish Cancel e Inthe File Format Options window ensure that the Same as input radio button is activated adjacent to Output file format This means the docking solutions will be written out in the same format as was used for input we saved our ligand out in MOL2 format thus this is the format our ligands will be written out in e Click on the button next to Output directory and specify a directory to which you have write permission this is where the GOLD output files will be written e Ensure that the Save rnk files Save ligand log files and Save initialised ligand files check boxes are switched on this will instruct GOLD to retain output files listing fitness function rankings and ligand log files The content of these files are discussed later see Analysis of Output e Click on the Information in File tab e Itis possible to write additional information to docked solution files This information is written to SD file tags for MOL2 files these tags are written to comment blocks This information is particularly important for post processing docking results with GoldMine For the purpose of this tutorial the nformation in File settings can be left at their default settings e Now click on the Selecting Solutions tab e GOLD can produce a large amoun
152. e e g hydroxyl and amino hydrogen atoms do not matter as they will be optimised during the GOLD run GOLD deduces hydrogen bonding abilities from the presence or absence of hydrogen atoms For example you can control the protonation state of a carboxylic acid group by adding or removing the ionisable hydrogen atom If incorrect ionisation or tautomeric states are inferred by the program it is unlikely that correct protein ligand binding modes will be predicted If you are unsure about e g the preferred ionisation state of the ligand you should perform separate GOLD runs using the different possibilities GOLD ignores atom charges both formal and partial It deduces whether an atom is charged by counting the bond orders of the bonds that it forms and comparing the result with the atom s normal valency GOLD User Guide 5 3 5 4 5 5 GOLD User Guide Ligand Geometry Conformation and Stereochemistry The ligand conformation will be varied by GOLD during docking The starting conformation therefore does not matter GOLD will not alter bond lengths or angles These parameters should therefore be set to reasonably optimum values A good practice is to build the ligand in an arbitrary conformation and then perform a few cycles of molecular mechanics minimisation to take the ligand close to its local potential energy minimum Ring conformations can be searched during docking using of library of ring templates see Ring Conform
153. e ASP file is quite strict incorrect editing may cause GOLD to behave in unexpected ways e The ASP fitness function shares many of its parameters with ChemScore see the chemscore params section see ChemScore for an explanation However please note that the default value of these may differ from the value used in ChemScore The asp paranms file e ASP COEFFICIENT Default 0 2 The total contribution to the score by the potential is scaled by a coefficient which has been optimised to 0 2 see The Generation of Potentials e CLASH COEFFICIENT Default 1 0 The CLASH COEFFICIENT controls the weight of the clash term to the overall score see The Generation of Potentials 72 GOLD User Guide e INTERNAL COEFFICIENT Default 1 0 The INTERNAL COEFFICIENT controls the weight of the internal energy of the ligand to the overall score see The Generation of Potentials e HBOND FUNCTION Default none The HBOND FUNCTION can be turned on by replacing NONE by ASP When switched on it will also include the metal correction see Metal and Hydrogen Bond Correction e HBOND CORRECTION FACTOR Default 1 0 see Metal and Hydrogen Bond Correction e CLASH FUNCTION Default ASP The CLASH FUNCTION is used for the calculation of clashes between the ligand and the protein The clash term is evaluated in the same way as for ChemScore see Clash Penalty and Internal
154. e Atom and Bond Types GOLD may be used in serial or parallel modes see Running in Parallel 2 2 1 Getting Started Overview of the GOLD Interface Select GOLD from the top level menu in the Hermes visualiser then Setup and Run a Docking from the resulting menu You will be asked whether you wish to create a new GOLD configuration file or to load an existing one The configuration file is a text file which specifies the GOLD calculation that is to be run including details of the ligand the protein binding site the fitness function to be used and the genetic algorithm parameters etc Selecting an existing configuration file e g from one of the tutorials will result in the defined configuration options being read into the GOLD Setup window The corresponding structure input files will also be opened within Hermes Selecting New will open an empty GOLD Setup window in which you will be required to specify all the configuration options required to define the docking job see Saving and Re using Program Settings in Configuration Files GOLD Setup mE GOLD has many configuration options which are available via the tree view at the left of this prita dialog To simp fy the process you can click the Wizard button below and you wil be guided Se through the essential configuration steps At the end of the wizard you can ether start the Lacking hice GOLD docking or access the more advanced configuration options Waters Ligand Flex
155. e Usage gold utils protonate i lt filename gt o lt filename gt rules lt filename gt e Details of the arguments above are protonate this is a required argument that instructs the script to protonate the supplied molecule file a this is a required argument that specifies the input molecule file o this is an optional argument that specifies the output molecule file rules this is an optional argument that forces the script to use the specified protonation rules file see Applying Protonation Rules 24 5 2 gold_utils print_rotamer e This utility is used for printing the rotamer block for the specified amino acid residue e Usage gold utils help print_rotamer i lt filename gt o lt filename gt residue lt residue id gt ignore_library reference ligand lt filename gt e Details of the arguments above are GOLD User Guide 231 print rotamer required argument that instructs the script to print he rotamer block for the specified amino acid residue for inclusion in a GOLD conf file i required argument that specifies the input molecule file o optional argument that specifies the output molecule file residue required argument that specifies the amino acid residue to use including the chain ID e g ASP102A ignore library optional argument that instructs the script to ignore the rotamer library and set all torsions to fully rotate reference ligand opti
156. e definition Activate the radio button that reads Ligand Choose the ligand you have just read in i e 1x0z ligand and select all atoms within 10 A of the ligand for the binding site definition This will make the binding site large enough to accommodate any GOLD User Guide possible binding mode of the PDESA inhibitor It can help here to switch off the display of H atoms using the Show hydrogens tick box in the top level menu of Hermes Carbon atoms outside of the binding site will turn purple Wizard step 3 Define the binding site The binding site can be defined by several different ways an atom a point or a reference ligand Atoms can be selected in the visualiser Global Options 119s 1TeF 1x0 2chm Wizard steps Point select atoms to define a centroid or edit XYZ 6 Choose a fitness function 7 GA search options 8 Finish 10 A V Detect cavity restrict atom selection to solvent accessible surface V Force all H bond donors acceptors to be treated as solvent accessible Add Definition as a Selection Click Next to proceed to the Optional load a configuration template dialogue At this point you are given the option to load a configuration file template Configuration templates can be used to load recommended settings for a number of different types of docking protocols In this example we will specify all docking settings manually Click Next to proceed to the Select ligands dialogue Selecting Ligands GO
157. e file rescore mol2 see Rescore settings 117 13 Docked Ligand Output Options 13 1 Specifying Ligand Solution File Formats and Directories e Click on Output Options from the list of Global Options given on the left of the GOLD Setup window then select the File Format Options tab e By default docked ligands will be written out in the same format as was used for input To change this specify the required file format by selecting either Same as input SD file or Mol2 e Use the Output directory entry box to specify the directory to which output files will be written or click on the button and use the directory selection window to choose the location When more than one ligand is being docked switch on the Create output sub directories check box if you want results for each ligand to be written to a separate sub directory gt GOLD Setup Mie E Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Wizard File Format Options Information in File Selecting Solutions Templates Proteins ai g a C ie C Define Binding Site Output file format Same as input SD file Mol2 Select Ligands fupt SS Waters Output directory output a Ligand Flexibility Fitness amp Search Options GA Settings J Create output sub directories For each ligand Output Options IV Save ligand rank rnk files
158. e fitness score is taken as the negative of the sum of the component energy terms so that larger fitness scores are better The external vdw score is multiplied by a factor of 1 375 when total fitness score is computed This is an empirical correction to encourage protein ligand hydrophobic contact During a docking run the fitness score may appear to get worse as the docking proceeds This is due to the fact that the effects of poor H bond geometry and close nonbonded contacts are artificially down weighted at early stages of the docking annealing Only the final fitness score i e from the completed docking has any meaning The fitness function has been optimised for the prediction of ligand binding positions rather than the prediction of binding affinities although some correlation with the latter has been found 59 7 3 2 Van der Waals and Hydrogen Bonding Annealing Parameters When GoldScore is being used the annealing parameters Van der Waals and Hydrogen Bonding allow poor hydrogen bonds to occur at the beginning of a genetic algorithm run in the expectation that they will evolve to better solutions At the start of a GOLD run external van der Waals vdw energies are cut off when E gt van der Waals kj where k is the depth of the vdw well between atoms i and j At the end of the run the cut off value is FINISH_VDW_LINEAR_CUTOFF This allows a few bad bumps to be tolerated at the beginning of the run Similarly t
159. e format SQlite is the default option Contact your database administrator for information on connecting to a PostgreSQL database e When using SQlite the GoldMine database filename to which the docked results will be saved should be specified Either enter the path and filename of the database file or click on the Browse button and use the file selection window to choose the file It will also be necessary to specify the name of the new dock set to which the poses need to be sent Alternatively results can be appended to an existing dock set Click on the Connect button and then select the required set from the Dock Set drop down list e Before running GOLD you must instruct GoldMine to receive docking results from the port specified To do this select GOLD from the top level menu within the Hermes visualiser then click on Receive Ligands from GOLD from the resulting drop down menu Alternatively this can be configured within the GoldMine Controller refer to the GoldMine user guide for further information e Click on Run GOLD The docking job will proceed and when complete the data should be saved in the appropriate GoldMine e Itis possible to have a GoldMine open whilst running the docking job It is also possible to take a selection of ligand poses from a GoldMine analysis and submit them to GOLD for docking see Receiving Ligands From GoldMine 136 GOLD User Guide 16 Saving and Reusing Docking Settings 16 1 Saving and Re using
160. e from the binding site have been deleted in order to speed up the calculation and hydrogen atoms have been placed on the protein in order to ensure that ionisation and tautomeric states are defined unambiguously see Essential Steps e The ligand from 1lpg has also been set up for docking see Essential Steps It is stored in 1 pg_ligand mol2 Again attention has been given to protonation states e g the benzamidine group has been built in its protonated form and the bond types have been set in accordance with GOLD conventions e These two files may be viewed in Hermes if desired Example conf Files and Output Files e Two GOLD configuration files have been prepared non_flexible conf this file was set up in the normal way using the GOLD front end It corresponds to a standard docking of the 1lgp ligand into the 1fax binding site using slow search settings 100 000 GA operations and allowing no side chain flexibility The considerations outlined in the preceding part of this tutorial suggest that this docking protocol is unlikely to give good results The corresponding output can be found in the non_flexible subdirectory 186 GOLD User Guide flexible conf this file defines a docking in which the Gln192 side chain is allowed to move It was set up using the Flexible Sidechains option in the GOLD front end The corresponding output can be found in the flexible subdirectory The processes used to setup and run these dockings a
161. e nitrogen is N am The atom type O co2 should be used for the oxygens of carboxylate and phosphate ions or the singly charged oxygen of phenolates If an atom is mis typed it is possible that GOLD will assign it the wrong H bond donor or acceptor properties Therefore correct atom type assignment is crucial An N 3 donor tetrahedral nitrogen is very different from an N 4 protonated nitrogen or an N p 3 planar trigonal nitrogen donor The assignment of rotatable bonds may also be affected If a bond has the wrong type it may be inappropriately allowed to rotate freely A list of atom and bond type conventions for some common difficult groups is available see Atom and Bond Type Conventions for Difficult Groups GOLD User Guide 6 4 Atom and Bond Type Conventions for Difficult Groups e Use of correct atom and bond types in GOLD is important for producing good results e In order for the GOLD atom type assigner to work correctly it is necessary for the input structures to have correct bond orders This can be difficult when a ligand contains a group that can be drawn in more than one way i e a group which has more than one canonical form In such cases there is usually a right and a wrong way for GOLD and you need to know which is which e The following table explains how to set the bond orders of some common difficult groups It also shows the atom types that GOLD will assign if bond types are set correctly or that you must
162. e of the Scoring function parameter file or click on the button and use the file selection window to choose the file e It was found necessary by Kirton et al to assign the planar nitrogens in the heme molecules as lipophilic when using the ChemScore scoring function In order to bring this about the chemscore p450 parameter files therefore contain the additional keyword MAKE PLANAR N LIPO 1 Note Use of this keyword has only been validated for nitrogen atoms within heme containing proteins Improvements in docking performance when used with non heme containing proteins are not guaranteed GOLD User Guide 8 Ligand Flexibility 8 1 Ring Conformations 8 1 1 Flipping Ring Corners To allow free corners of ligand rings to flip during docking click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window activate the flip ring corners tick box in the Explore ring conformations section of the window This will result in GOLD performing a limited conformational search of cyclic systems by allowing free corners of rings to flip above or below the plane of their neighbouring atoms The rules governing flipping of ring corners in GOLD are given in A W R Payne and R C Glen J Mol Graphics 10 74 91 1993 8 1 2 Using CSD Ring Conformation Templates A library of ring conformations extracted from the Cambridge Structural Database CSD can be utilised by GOLD This allows GOLD
163. eaaseeeeeseceeeas 143 20 1 Tutorial 1 A Step By Step Guide to Using GOLD cececcceccceeeeseeeseeeeees 143 20 2 Tutorial 2 Handling of Metals in GOLD cc ceeeccceccececeseeeseeeeeeeeeesaaaseeeeeees 157 20 3 Tutorial 3 Use of Hydrogen Bonding Constraints cccceccecceeeeseeseeeeeeees 164 20 4 Tutorial 4 Use of Substructure Based Distance Constraints 172 20 5 Tutorial 5 Docking with Water in the Binding Site cccccceccceesssesseeeeeeees 179 20 6 Tutorial 6 Docking with a Flexible Side Chain ccccccssssssssecceeeeeeeesseseeeees 185 20 7 Tutorial 7 Docking using Localised Soft Potentials cccccccccccceessssseeeeeees 194 20 8 Tutorial 8 Generating Diverse Solutions cccecccccccccceeeeeeeeeceeeeesssaaseeeeess 198 20 9 Tutorial 9 Running a Covalent Docking ccesccccccceecceseseeeeeeeeesseaaeseeeeeess 203 20 10 Tutorial 10 Ensemble Docking ccccccccccccasseseeeeeeeeecssaaeseeeeeeessuaaaeseeeeess 209 Appendix B List of Atom and Bond TyPe s ccccccsssssssseeccceeesaeeeeeeeeeeeessaeeeseeeeeeeeeeaa 220 Appendix C Additional Tags in Output FileS cccecccccccccceceeeseeeeceeeeeseaeeenseeeeeeeeeeas 221 Appendix D Genetic Algorithm Parameter Definitions ccccccccccesecsssseeeeeeeeeeeeaes 226 2321 POPULATION Size fev vecceesceccccecatavedesceecatatanseesssetesc datas ssedsdehesetabnsesedstedetetnandesebeds 226 23 22 Selection Pr S
164. eaning Name in ChemScore File Default Value The ideal H A X angle in degrees BETA IDEAL 180 0 The absolute deviation of the actual H A X Calculated for each H bond angle from 8 The tolerance window around the H A X DELTA BETA IDEAL 70 0 angle within which the H bond is regarded as ideal The maximum possible deviation from the DELTA_BETA_MAX 80 0 ideal H A X angle above this the interaction is not regarded as an H bond The Gaussian smearing sigma associated with HBOND BETA SIGMA 10 0 this term The third block function in the H bond equation B is the sum of all possible values for a given hydrogen bond For example a tertiary amine acceptor has three covalently bound atoms that could be deemed as the X atom in this case the term added for an H bond to the amine is the product of the block function values for all three possible H A X angles Hydrogen bonds have a regression coefficient associated with them v By default this is set to 3 34 The name of this coefficient in the ChemScore parameter file GOLD User Guide see Altering ChemScore Fitness Function Parameters the ChemScore File is HBOND_ COEFFICIENT 7 4 4 Metal Binding and Lipophilic Terms e The metal binding term in ChemScore is computed as a sum over all possible metal ion acceptor pairs where the acceptor is an atom in the ligand that is capable of binding to a metal e Each term in the summation is
165. eck the quality of the input ligand structure e check _mol2 can either be used to simply generate output on the ligand of interest or to create a corrected version of the ligand that can be used by GOLD e check _mol2 only works on mol2 files with individual ligands In other words it will not work on a multi mol2 file e check_mol2 can be invoked from the command line The following platform dependent commands should be used Linux platforms lt install dir gt GOLD Suite bin check mol2 i ligand mol2 230 GOLD User Guide o corrected _ligand mol2 Windows platforms at the Windows command prompt lt install dir gt GOLD gold d_win32 bin check mol2 exe i ligand mol2 o corrected _ligand mol2 where lt install_dir gt is the GOLD installation directory If specifying the full path the command will need to be in inverted commas e g C Program Files CCDC GOLD_Suite GOLD gold d_win32 bin check mol2 ex e i ligand mol2 o corrected _ligand mol2 e The flags are original input ligand o corrected output ligand 24 5 gold_utils e The gold_utils file contains four scripts that allow modifications to be made to input and or output files thus facilitating docking setup and post processing On Windows machines this file is located in C Program Files x86 CCDC goldsuite 5 3 Hermes gold_utils exe 24 5 1 gold_utils protonate e This utility is used for protonating molecule file s
166. ection window to choose the file The parameters read in from the configuration file will overwrite any parameters that have already been set in the GOLD front end If you have a valid configuration file i e one that completely specifies a GOLD job you can run GOLD from the command line by using a simple command available in GOLD_DIR bin For example if the configuration file is gold conf the command is gold auto gold conf amp 16 2 Using Configuration File Templates GOLD User Guide The configuration file is a text file which fully specifies the GOLD job that is to be run Once a configuration file has been created it can be saved and re used either as a quick way of reading program settings into the GOLD front end or to run GOLD from the command line see Saving and Re using Program Settings in Configuration Files Configuration file templates can be used These contain recommended setting for a number of different docking protocols Recent validation experiments carried out on the DUD sets of actives decoys have suggested preferred virtual screening protocols for several protein target classes These protocols show good early enrichment statistics but also have the best trade off between speed and accuracy as far as can be ascertained New protocols for nuclear hormone receptors kinases metallo proteases and folate containing 137 enzymes have been created The protocol for serine proteases has been changed to allo
167. ed control can be achieved by using the rotatable bond override mol2 file found in the SGOLD_DIR gold directory Some fragments are already provided which can be edited however user specific ones may also be added Instructions on how to do this as well as further information can be found in the file itself To post process fragments via the rotatable bond_override file click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Postprocess Rotatable Bonds check box Then either enter the path and filename of the file or click on the button and use the file selection window to choose the file This option is particularly useful if further control is sought over more than one ligand with a common substructure in a ligand library file The new bond type s are specified in the rotatable_bond_override mol2 file in the lt TRIPOS gt COMMENT part of the molecule file The following format should be used RESET BOND TYPE lt bond_number gt lt fix flip 1 am fix keeps the bond at its input angle This option can also be specified for a single ligand docking via the gold conf see Fixing Rotatable Bonds at Their Input Conformation flip causes 180 degree turns of the input angle geometry 1 re types the bond to a single bond thus it is treated as fully rotatable amre types the bond as an amide bond A report detailing what has been ma
168. ed in the Hermes User Guide 3 6 7 Solvent Accessibility GOLD User Guide Each atom in the defined binding site is tested for solvent accessibility this is a two step process First the solvent accessible surface of each atom in the defined binding site is calculated Potential donor and acceptor fitting points used for ligand placement are then generated for only those protein atoms that are accessible Second the potential fitting points are themselves tested for solvent accessibility and only those fitting points that are accessible are used It is possible to remove this requirement for fitting points to be solvent accessible In this case fitting points would be generated for all solvent accessible donor and acceptor atoms within the binding site Remember that these atoms are already deemed to be solvent accessible but it s their potential fitting points that may have been desolvated by neighbouring atoms This option can be used e g to avoid problems with solvent accessibility of backbone carbonyls in kinases where one of the carbonyl lone pairs is typically desolvated by a neighbouring atom To generate fitting points for all solvent accessible donor and acceptor atoms switch on the check box labelled Force all Hbond donors acceptors to be treated as solvent accessible This option is available by clicking on Define Binding Site from the list of Global Options given on the left of the GOLD Setup windo
169. ed position i e expressed with respect to the same coordinate frame as the protein and with the coordinates required to place it in the correct pose To specify the template file either enter the path and filename of the file or click on the Template File button and use the file selection window to choose the file GOLD User Guide e The weight term determines the maximum energy term that would be added to the score in the case of perfect overlap between ligand and template As an initial value for this term we suggest a value between 5 and 30 3 GOLD Setup BE x Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial gold conf Load Save Options Protein 1cil lyase oxo acid Wizard Cc Fitness amp Search Options Shape overlap GA Settings Output Options Template file fnd Settings henderson Desktop gold_tutorials tutorial4 template mol2 GoldMine Parallel GOLD i i l Constraints Constraint weight 10 0 Similarity Scaffold Region Atom Typing Templates Type of similarity Proteins Define Binding Site C H bond donor overlap Select Ligands waters H bond acceptor overlap Ligand Flexibility J Never dock a ligand when a constraint is physically impossible Help R Run GOLD Run GOLD In The Background Finish Cancel e Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor It is po
170. ediating hydrogen bonds between protein and ligand or be displaced by the ligand on binding GOLD allows waters to switch on and off i e to be bound or displaced and to rotate around their three principal axes to optimise hydrogen bonding during docking To predict whether a specific water molecule should be bound or displaced GOLD estimates the free energy change AG associated with transferring a water molecule from the bulk solvent to its binding site in a protein ligand complex AG for a given water molecule is defined as AG W AG W AG W AG W is a constant penalty added for each water molecule that is switched on and represents the loss of rigid body entropy on binding to the target hence rewarding water displacement Note AG values were optimised against a training set of 58 protein ligand complexes for four targets HIV 1 protease factor Xa thymidine kinase and the oligopeptide binding protein Opp A where water molecule play key roles in the recognition Further details can be found in Modelling Water Molecules in Protein Ligand Docking Using GOLD see References AG W represents the intrinsic binding affinity of a water molecule and contains contributions resulting from interactions that the water forms with the protein and ligand changes in the interactions between protein and ligand caused by introduction of the water are also accounted for Therefore for a water molecule to be bound to a protein lig
171. eeeeeeeseeaeaseeeeeeeeesaeaaesss 100 10 5 1 Method Used for Similarity Constraints ccccccccssssssseeeeeeeeeeeas 100 10 5 2 Setting Up a Similarity Constraint cc ccccccccccceceeeesesseeeeeeeeeeaes 100 10 6 Scaffold Match Constraint e ec ccccceeeeeeesnneeeeeeeeeeeeenaaeeeeeeeeeeeeeaaaeeeeeeeeeeeea 101 10 6 1 Method Used for Scaffold Match Constraint eese 101 10 6 2 Setting Up Scaffold Match Constraints cccccccccccscssesssseeeeeeeeeeaes 102 10 7 Interaction Motif Constraint 0 0 eeeceeeccceeeeeeeeeeeeenaaeeeeeeeeeeeeeaaaeeeeeeeeeeees 103 10 7 1 Method Used for the Interaction Motif Constraint cece 103 10 7 2 Setting up an Interaction Motif Constraint ccccccsssseeeeeeeeeeeees 104 Balancing Docking Accuracy and Speed cccccssssssseeeccceecaeseseecccecessaeeeeeeeeeeeesaaaagess 107 It Numberot DOckings 2 n08 28 nGnswe 2 Sen ee ON eee 107 TEZ Early Termination siere ea secede rE EE TEKEE E TEPEE EEEE Eaa 107 11 3 Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 108 11 3 1 Genetic Algorithm Overview cccccccccccccsssseseeeeeeeeeeaeaeeeeeeeeeeeaaa 108 11 3 2 Relationship between Genetic Algorithm Parameters and Speed 109 GOLD User Guide 11 3 3 Using Automatic Ligand Dependent Genetic Algorithm Parameter SO tliN r E 110 11 3 4 Using Preset Genetic Algorithm Parameter SettingsS ccccc0008 112 11 3 5 Using User Defined
172. eeees 45 5 7 3 Setting Up Substructure Based Covalent LINKS cccccsssssseeeeeeees 46 5 8 Specifying a Ligand Reference File cccccccccssssseseeecceeeceaeesseeeceeessausanseseeees 47 Atom and BOnd TYPOS cccccsessesecccccccceeeseeeeceeeesaeeeseeeeeeeesseaeeeseeeeeeesseaaaaeeeeeeeesaaaaaess 49 6 1 Atom and Bond Type Overview cccccccccccsssssseseceececsaeeesseeceeeeesaaaeaseeseeeeeeaaa 49 6 2 Automatically Setting Atom and Bond Typ S ccccccsssssesseeccceeeeeeaeeseeseeeeeeaas 49 6 3 Manually Setting Atom and Bond Types ccccccccccsesssseeeeeeeeesaaeeaeseseeeeeeeaas 50 6 4 Atom and Bond Type Conventions for Difficult GroupS ccsssseeeeeeeeeeeees 51 6 5 Internal GOLD Atom Ty pesiaancannnnnncn ened Ae aaad HAAR A AAA AAR 53 FIMESS PUNCH ONS gt cccstsoctetst estetete nRa RANAN RRON ONSON NN ONONO NNNUNN OONN EN 54 7 1 Selecting a Fitness Function cccccccsssssseeccceecceeeesesecceeesesaaeseeseeeeeesaaaaasesseees 54 7 2 Piecewise Linear Potential CHEMPLP cccccccccccceeeeeeeeeeeeeeeeseeeeeeeeeeeeeeeeess 54 7 2 1 OVERVIEW 5 oe a Roscoe te eee ee ween ROA AAA POr RA bene bebop se RA PATET 54 7 2 2 PLP Interaction TYP S cccccssseccccssseccceeeseeceeeeseeeeeaeseesseeesessaeeseeees 55 7 2 3 Altering PLP Fitness Function parameters ccccccccccsssssseeeeeeeeeeees 56 7 3 GOldSCOMre nse E EE E E O aan 59 7 3 1 OVERVIEW Sorsrsrrts rarita toras E tat S ETO
173. eeeesseaeesseeeeeeessaaaagess 124 14 5 1 Using GOLD with a Grid Engine environment ccceeceeeeeeeeeeees 124 14 5 2 Using GOLD with Parallel Virtual Machine PVM cccceceeeeeeees 124 15 Viewing and Analysing R SUItS ccccccssssssscccecccceeeesseeeeeeeeesaseeeseseeeeeessaeaaeeseeeeeesaaa 126 15 1 Description OF OUtPUt FIGS ss ccceceseseeeseeeseeayes sevesesesteesveedevisesteeeeesdeets steers 126 15 1 1 Files Containing the Initialised Protein and Ligand ccccccc0 126 15 1 2 Files Containing the Docked Ligand s ccccccccccccccceeeeeeeeeeeeeeeees 126 15 1 3 File Containing the Protein Binding Site Geometry cccccceeeee 127 15 1 4 File Containing Ranked Fitness Scores for an Individual Ligand 127 15 1 5 File Containing Ranked Fitness Scores for a Set of Ligands 128 15 1 6 RescoreSolution File s2a 224nacansndn ean Hanae 128 15 47 RESCOME L g Filiera eee e eee ae oe eee ee on 129 15 1 8 Protein Log File eeccccccecccccesesseseeeeeeesseeeeseeeeeeeeeseaaeseeeeeeeeeaaa 129 15 1 9 Ligand Log File cc ceecccccccccccceeesseeeeceeeeeaeeeeseeeeeeeeeseaaaseeeeeeeeeeaaa 130 15 1 10 File Containing Error Messages cccccccccesssseeeeeceeeeseeeesseeeeeeeeeaas 130 15 1 11 Process File eee ccessccneeeeeeeeeeeeeaaeeeeeeeeeeeeesaaaaeeeeeeeeeetsecaeeeeeees 130 15 1127 Seed log Filenin na eer 131 15 2 Information on the Progress of Docking R
174. eeeessuaaaeeeeeeeeesaaa 80 8 3 Flipping Pyramidal NitrOgens cccccccccsssssssseeccccecesaaessseececeeesauaeaeeeeeeeeeeaas 80 8 4 Intramolecular Hydrogen BONds ccccccccsssssseeeccceeecsaeeeeeececeeesaaaaeeseseceeeesaaa 80 8 5 Flipping Planar NitrOgens ccccceesssecccccccceseeeeeecccceeseseeseeeeeeeessssaaaeseseeeeeeeaaa 81 8 6 Protonated CarboxyliG Acids ennei iieii ii 81 8 7 Using Torsion Angle DistribUtions ccccccssseseecccccecceeeseeecceeeesaeeaseeeeeeeeesaas 81 GOLD User Guide v vi 10 11 8 7 1 Enabling Use of Torsion Angle Distributions ccccccseceeeeseeeeeees 81 8 7 2 Editing Torsion Angle Distribution Files ccccccsssssseseeeeeeeeeeeeeeees 82 8 7 3 Matching Torsion Angle Distributions at RUN Time ccccceseeeeeees 82 8 8 Overriding Automatic Bond Settings cccccccsssssssseeccceecseeesseeeeceeeesaeaaseeseeees 83 8 9 Fixing Rotatable Bonds at Their Input Conformation cccccccsssssseeseeeeeeeeaes 84 Ligand Search Options cccccccccccccccaeessseeececeecsaeeesseeeeeeeeseeeaeseeeeeeeeessaaaaseeeeeeeeesaaagaess 86 9 1 Internal EnereyiOttset icccccosevecececacececevevectetcteteeces weedetes ccececes weedeeee cceteees seeteee tee 86 9 2 Hydrophobic Fitting Points cccccccccccccccseeesseeeeceeeeesaeeesseeceeeessseaeaeeeseeeeeeeaas 86 9 3 Generating Diverse SOIUtIONS ccccceseseseccccccceaeseseecceeeesaaueeseeeeeeeeesauaae
175. een able to optimise their interactions with H bond acceptor functionality in the protein This 183 means that both are making three good H bonds The third water has been excluded in all the reported docking poses Changing the Scoring function e You may wish to stop the tutorial here However optionally you can run through the tutorial again this time having either having the CHEMPLP GoldScore or ASP option set in Fitness amp Search Options in the GOLD GUI e You will find that similar results are obtained When all waters are turned off two binding modes are generally found that score similarly well One of these binding modes actually superimposes the reference ligand very well and allowing waters to toggle does not significantly improved the superimposition in this case However allowing the waters to toggle does result in only this one binding mode being returned The second spurious binding mode is successfully eliminated This ends the tutorial 184 GOLD User Guide 20 6 Tutorial 6 Docking with a Flexible Side Chain Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial6 toa directory to which you have write permissions The object of this tutorial is to demonstrate how to dock a ligand into a binding site which is known to contain a flexible side chain The example will involve docking the ligand from PDB entry 1lpg into the protein binding site taken from 1fax These
176. een written to the output directory each containing the results of a separate docking attempt e The result of each docking attempt is written out as gold_soln_ligand_m1_n mol2 where n is the number of the docking solution 1 2 3 and m1 is an index to the ligand in this example only one ligand was docked e Note that the file go d_so ln_ligand_m1_1 mol2 is not the best GOLD prediction it is just the solution found in the first docking attempt However as GOLD proceeds symbolic links are created ranked_ligand_m1_1 mol2 will point to the current top ranked solution ranked_ligand_m1_2 mol2 will point to the second best solution and so on e Return to the Hermes 3D view and inspect the top ranked solution predicted by GOLD Note that the original protein we edited is still loaded to make the display less complicated you may wish to disable one of the proteins by deactivating the tickbox GOLD User Guide 155 adjacent to 1ACM or 1ACM_2 under the Display tab in the Molecule Explorer If you do this return to the Docking Solutions tab once you have finished e The docking solutions are given in their docked order with their corresponding fitness score listed under the column headed PLP Fitness If required the solutions can be ordered by clicking on PLP Fitness to determine which is the highest scoring e Asimple test of the effectiveness of a docking program is to take a protein ligand complex from the PDB and extract the ligand
177. eight is a user defined score that determines how good a hydrogen bonding interaction has to be in order for it to be considered a hydrogen bond by GOLD The Minimum H bond geometry weight takes a range of values from 0 to 1 by default this value is set at 0 005 Specifying Multiple Constraints 166 It is possible to specify several different protein H bond constraints with different weights for each constraint Simply select each protein atom required to form an H bond with the ligand as well as the required weight then click on the Add button to add the constraint definition to the constraints dialogue at the bottom of the GOLD User Guide window Repeat the procedure to set up further constraints each constraint will be displayed on a separate line in the constraints dialogue e For a given protein H bond constraint more than one protein atom can be selected and added to the Protein atom s required to form H bond input box This will instruct GOLD to use an either or type of constraint during docking For example specifying two protein atoms m and n separated by a space will result in the constraint being satisfied if an H bond is formed to either m or n during docking This is of particular use when defining constraints involving for example carboxylates where it is not important which oxygen atom forms an H bond provided one does Defining the Protein H Bond Constraints e The crystal structures of HIV 1 protease in comp
178. electing Load Existing from the resultant pop up window navigating to the folder to which you copied the tutorials files selecting the file gold conf and clicking Open We will need to identify the waters in the binding site that we particularly want to consider and set up their chosen states To do this pick Configure Waters from the list of available options The dialogue is empty so we need to specify our water molecules by reading in the water files To do this click on the Add button select the three water files then hit Open The water molecules will be listed in the Configure Waters dialogue By default the waters are allowed to toggle and spin This sets up a water so that it may either be removed or kept and made use of in terms of hydrogen bonding depending on which arrangement scores most highly for a given ligand pose On sets the water to be always present in the binding site and allows the hydrogen positions to vary during docking in order to maximise the hydrogen bonding score both from interactions with the protein and the ligand The Off water state option allows a water to be removed from consideration during docking We will be using both these options shortly GOLD User Guide 3 GOLD Setup BBE Conf File C Program Files C OLD Suite GOLD examples tutorialS gold conf Global Options Protein 1acj hydrolase carboxylic esterase Wizard Templates index toggle state spin st Proteins Define
179. emble The best ligand conformation found in any of the ensemble structures is returned i e GOLD selects the best protein for a particular ligand based on the maximum fitness value of a ligand For example if a ligand gets the scores 10 in protein 1 20 in protein 2 and 15 in protein 3 protein 2 will be selected There should only be one binding site definition across the entire ensemble hence the need to superimpose proteins and this must be protein independent 4 3 2 Setting up Proteins for Ensemble Docking GOLD User Guide Proteins being specified in an ensemble should be set up in the usual way see Setting Up the Protein s In addition proteins that are to be used in an ensemble docking must be superimposed 33 Proteins can be superimposed by using the Superimpose button in the Proteins window of the GOLD interface or via Calculate and then Superimpose Proteins in Hermes Brief details follow complete details are provided in the Hermes documentation Search for sequence alignment tool x Superimposing proteins can optionally use sequence alignment For the purposes of pre 7 defining which residues in one protein align to other residues in a second protein This can be done using ggsearch32 a component of the fasta package available from http Fasta bioch virginia edujFasta_www2 Fasta_down shtml Would you like to search For the appropriate binary on your computer now Yes No A wizard is provided to
180. en the ligand being docked and the template provided The similarity between the two is evaluated as a Gaussian overlap term The similarity constraint can be applied in three ways that differ in the way that the overlap between ligand and template is calculated The similarity can be evaluated by using the overlap between all donor atoms in the template and the ligand being docked by using the overlap between all acceptor atoms in the template and the ligand being docked by using the overlap of all atoms of the template this can be regarded as a ligand shape constraint The energy term to be added is calculated as similarity times weight the similarity value is between 0 and 1 where 1 indicates identity of template and ligand If you wish to place a fragment at an exact specified position in the binding site as opposed to biasing the docking use the scaffold match constraint see Scaffold Match Constraint Setting Up a Similarity Constraint To define a similarity constraint click on Similarity from the list of Global Options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options Specify the similarity type to be used by selecting H bond donor overlap H bond acceptor overlap or shape overlap see Method Used for Similarity Constraints The similarity template file should contain the template molecule or fragment in its dock
181. ensive functionality for setting up protein and ligand files see Essential Steps for setting up the protein file and Essential Steps for setting up ligand files we recommend you use a molecular modelling program Full details of the software requirements needed in order to use GOLD are given elsewhere see Introduction Please note Due to the non deterministic nature of GOLD results may vary from those described in the tutorials 20 1 Tutorial 1 A Step By Step Guide to Using GOLD Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutoriall toa directory to which you have write permissions GOLD features a Wizard for docking setup and an Advanced interface for users who are more familiar with using GOLD The Wizard guides the user through the key steps involved in setting up protein and ligand files as well as the components that are key to running a successful docking GOLD will only produce reliable results if the protein and ligand input files are set up correctly It is therefore essential that a number of key steps are followed when preparing any input structure for use in GOLD see Essential Steps for setting up the protein file and Essential Steps for setting up the ligand files This tutorial aims to provide a step by step guide to making the most of the GOLD wizard To illustrate this the procedure for setting up a protein and ligand for use with GOLD then the subsequent docking will be
182. entifies each ring in the ligand and attempts to match it to a ring template in the following file lt GOLD DIR gt ring conformations template library mol2 77 8 1 4 78 Note that the atom types in the template _library mo12 file must match the ligand atom types exactly i e after any ligand atom typing has been performed see Automatically Setting Atom and Bond Types Details of matched rings are written to the gold lt ligand_ name gt m lt n gt 1log file under the heading Match Ring Templates The template library mol2 file contains 1274 ring templates Each template represents a different ring identified within the Cambridge Structural Database CSD For each template the number of alternative conformations that will be explored during docking will vary depending on the abundance and suitability of data within the CSD The index number of the final ring conformation used in each docking solution is written to the gold_ lt ligand_name gt m lt n gt 1log file under the heading Chromosome decoded User Defined Ring Conformations GOLD can vary ligand ring conformations during docking A library of ring conformations extracted from the Cambridge Structural Database CSD is supplied with GOLD for this purpose see The CSD Ring Conformation Library and Matching Templates at Run Time It is also possible to specify your own ring templates and the allowed alternative conformations for those rings This is useful if you wish to
183. entry code 1qbt The use of hydrogen bonding constraints in order to reproduce these key interactions will also be illustrated Open Hermes and read in and inspect the file protein mol2 from the folder to which you copied the tutorial3 files The original PDB file 1QBT pdb has also been provided should you wish to set up the protein for yourself HIV 1 protease protein mol2 has already been set up in accordance with the guidelines for the preparation of protein input files see Setting Up the Protein s An important feature of cyclic urea inhibitors is their ability upon binding to displace a structural water molecule present within the active site of the protein In this example all water molecules have been deleted from protein mol2 However in other complexes you may not know whether water molecules should form mediating hydrogen bonds or be displaced by the ligand on binding GOLD allows waters to switch on and off i e to be bound or displaced to rotate and to translate within a radius of 2A to optimise hydrogen bonding during docking see Water Molecules The cyclic urea inhibitor has already been prepared in accordance with the requirements for setting up the ligand see Setting Up Ligands Open the file igand mol2 from the folder to which you copied the tutoria13 files within Hermes and inspect the structure Keep the file open once you have finished A configuration file has been provided for this tutorial The gold con
184. er To protonate the ND1 and or NE2 atoms enable the corresponding check box es and click on the Set Protonation button e If you are unsure about the tautomeric state of a His residue you should perform separate GOLD runs using the different possibilities 3 4 Deleting Ligands and Metal lons e The protein file may have one or more ligands occupying the binding site that must be removed before you can perform a docking e The removal of ligands is protein specific thus first select the appropriate protein tab adjacent to the Global Options tab then click on Delete Ligands from the list of options given e A list of the ligands present in the protein file will be displayed Each ligand is assigned a unique identifier based on the protein chain 3 GOLD Setup Bik Conf file ee eel load Save Options 1ACM 1a42 187 Extract and Reload Protonation amp Tautomers Flexible Sidechains Soft Potentials Metals Constraints Covalent Interaction Motif Extract Your protein may have one or more ligands occupying the binding site that must be removed before you can peform a docking Extracted ligands are automatically reloaded so they can be used to define the binding site and are written to file for later comparision with docking results Help R Run GOLD Run GOLD In The Background Finish Cancel e Clicking on a ligand in this list will highlight it in
185. er the water should be present or absent i e bound or displaced by the ligand during docking The orientation of the water hydrogen atoms available options are Spin have GOLD automatically optimise the orientation of the hydrogen atoms GOLD User Guide 11 3 6 3 6 1 12 Trans_spin activate this option and input a translation value into the distance dialogue to make GOLD spin and translate the water molecule to optimise the orientation of the hydrogen atoms as well as the water molecule s position within a user defined radius Note that the distance value must be between 0 and 2 A Fix use the orientation specified in the input file After docking a summary of which waters were retained or displaced and their contribution to the fitness score can be found in the Analysis of active water placements section of the gold_ligand 1og file Defining the Binding Site Overview It is necessary to define the protein binding site This can be done in several ways e g by specifying the approximate centre of the binding site and taking all atoms that lie within a specified radius of this point see Defining a Binding Site from a Point The binding site definition is detailed in the Cavity atoms section of the gold_protein log file The cavity atom selection can be saved as a protein atom subset and viewed within Hermes To do this click on the Add Definition as a Selection button within the Define Binding Site section of the GOLD
186. es Containing the Initialised Protein and Ligand gold_ligand mo12 is the initialised ligand datafile with lone pairs added and the sets DONOR_HYDROGENS and LONE_ PAIRS defined If you do not wish to save this file click on Output Options from the list of Global Options given on the left of the GOLD Setup window select the File Format Options tab then disable the Save initialised ligand files check box gold_protein mo12 is the initialised protein datafile with lone pairs added to binding site atoms and the sets DONOR_HYDROGENS and LONE PAIRS defined The binding site is defined in the set CAVITY ATOMS These set definitions in the gold _protein mo12 file are available for visualisation as protein subsets in Hermes Files Containing the Docked Ligand s Each ligand will normally be docked several times so a given input ligand will produce a set of files each containing the results of a separate docking attempt Suppose that the original ligand file is structure mo12 this can contain more than one ligand in which case each will be docked As the GOLD job progresses the result of each docking attempt is written out as gold soln structure m n mol2 where nis the solution number 1 2 3 and m is the number of the ligand i e m1 for the first ligand m2 for the second etc Note that the file gold_soln_ structure m1_1 mo12 is not the best GOLD prediction it is just the solution found in the first docking attempt However a
187. eseeess 87 9 3 1 Method Used to Generate Diverse Solutions 87 9 3 2 Setting Up GOLD to Generate Diverse Solutions ccccsssseeeeeeeees 88 Setting CONStraINtS ceeccccssseccccceeececcaeeecccseusecenseueeessaueeceeseuseeeeseueeesseaeeessuasseesaaases 89 10 1 Using the Constraint Editor ccccccccssssssseecceceeeeeeeeseeeceeeeseeaeeeseeeeeesessaaaaness 89 10 2 Distance COnstrai nts sc cccicceicteupseacear anaran Tana Tana eases Sana ana teases toactea teaches tocaceey 91 10 2 1 Setting Up a Distance Constraint cccccccssseececceeeeeeeeeeeeessaeeeeeees 92 10 2 2 Method Used for Substructure Based Distance Constraints 93 10 2 3 Setting Up Substructure Based Distance Constraints ccccccc 94 10 3 Hydrogen Bond Constraints cccccccccsssssseeecceccecaeaeesseecceeeessaaasseeeeeeeessuaaggsss 95 10 3 1 Setting Up Hydrogen Bond Constraints ccccccccceesssssseeeeeeeeeeeees 96 10 3 2 Method Used for Protein H Bond Constraints 96 10 3 3 Setting up Protein H Bond Constraints cccccccccccessassseeeeeeeeeeeaes 97 10 4 Region Hydrophobic Constraints cccccccssssssssssssssssesssssssessssesssssseesseesssaes 98 10 4 1 Method Used for Region Hydrophobic Constraints cccceeeees 98 10 4 2 Setting Up Region Hydrophobic Constraints ccccccccccccceseeeeeeees 98 10 5 Similarity Constraints cccccccceccccccccceeesesececceeeceaaeese
188. eter file is stored in the GOLD distribution directory It contains all the parameters used by the GOLD implementation of ChemScore A full description of the meaning of the various parameters is given elsewhere see ChemScore 68 GOLD User Guide 7 5 7 5 1 7 5 2 GOLD User Guide The ChemScore file can be customised by copying it editing the copy and instructing GOLD to use the edited file To use a modified chemscore params file click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select ChemScore from the Scoring Function drop down menu Then either enter the path and filename of the Scoring function parameter file or click on the button and use the file selection window to choose the file The format of the ChemScore file is quite strict incorrect editing may cause GOLD to behave in unexpected ways or even to crash Because of the large number of parameters no guarantee can be given that the program will behave reliably with anything other than the default parameterisation Astex Statistical Potential ASP Overview Th iv For a more thorough discussion on the Astex Scoring Potential ASP fitness function please see W T M Mooij and M L Verdonk Proteins Struct Func And Bioinf 61 272 287 2005 ASP is an atom atom potential derived from a database of protein ligand complexes and can be compared to other such scoring potentials e g P
189. ettings are encapsulated into three speeds Slow most accurate this equates to 100 000 GA operations Medium 50 000 operations Fast least accurate 10 000 operations e There is a trade off between speed and reliability The fewer operations the faster the docking but the search space will be less well explored e Further options are available by clicking on the More gt gt button Enable automatic GA settings by clicking on the Automatic radio button and ensure the Search efficiency is set to 100 This will make GOLD automatically calculate an optimal number of operations for a given ligand thereby making the most efficient use of search time e g small ligands containing only one or two rotatable bonds will generally require fewer genetic operations than larger highly flexible ligands GOLD User Guide 215 Wizard step 7 Genetic Algorithm search options The time taken to perform a docking is a balance between speed and accuracy the slower the docking the more accurate it will be GOLD Setup Global Options ITBF Wizard steps 1 Select a protein 2 Protein setup 3 Define the binding site 4 Configuration template 5 Select ligands 6 Choose a fitness function 7 GA search options 8 Finish 1t9s 1xp0 2chm Automatic gt Preset User defined Search efficiency C Min ops 10000 Library Screening Virtual Screening Hit Next to proceed to the Finish basic GOLD co
190. etween different tautomeric states during docking Delete Waters see Deleting Waters water molecules often play key roles in protein ligand recognition Water molecules can either form mediating hydrogen bonds between protein and ligand or be displaced by the ligand on binding Water molecules within the active site can be retained and allowed to toggle i e switch on and off during docking rotate and translate within a radius of 2A to optimise their H bonding positions Those outside of the binding site can be removed from the protein altogether Delete ligands see Extracting and Deleting Ligands the 1ACM pdb protein is the raw PDB file which is the original protein ligand complex For GOLD to effectively dock a ligand back into the active site the co crystallised ligand must first be removed Adding Hydrogen Atoms From within the JACM tab add hydrogens to the protein by selecting the Add Hydrogens button from the first Protonation amp Tautomers option Note that this may take a little time depending on how many other processes are running on your computer Once the H atoms have been added a pop up window will inform you that 7196 H atoms were added Click OK to close the pop up window Deleting Waters Still in the JACM window hit the Extract Delete Waters option underneath Protonation amp Tautomers From within this dialogue it is possible to specify water molecules that mediate protein ligand interactions i e act
191. explained and additional information will be provided on related issues In this example GOLD will be used to determine the binding mode of N phosphonacetyl L aspartate with the aspartate carbamoyltransferase PDB entry code 1acm Using the GOLD Wizard to Prepare the Protein File GOLD User Guide Open Hermes Open the GOLD setup wizard by clicking on the main menu option GOLD then by picking Wizard from the resultant pull down menu The steps required to setup files for docking are listed down the left hand side 143 i GOLD Setup 5 Wizard step 1 Select one or more proteins Either choose a protein already loaded in the visualiser or load a new file Global Options aparin Select proteins to use Load Protein Superimpose Proteins 2 Protein setup 7 3 Define the binding site 4 Configuration template 5 Select ligands 6 Choose a fitness function 7 GA search options 8 Finish List all loaded files not just proteins Protein score offset ensemble docking only Protein Score Offset Fix all protein rotatable bonds Help R Back Next gt Cancel Wizard Selecting a Protein 144 In the Select proteins to use window read in the protein file 1ACM pdb by hitting the Load Protein button navigating to the folder to where you copied the tutoriall files select the file and then clicking on Open The protein file will be loaded into Hermes 3D view You will
192. f is loaded by clicking on the main menu option GOLD then Setup and Run a Docking From the resultant pop up window select the Load Existing button you should then browse to the directory to which you copied the tutorial3 files select the file gold conf then hit Open This will automatically load the settings and parameter values for this tutorial into the GOLD front end in addition to the specified protein file The GOLD interface contains two tabbed views the default is Global Options which allows you to specify particulars of the docking in general the other displays the protein name in this case Protein 1qbt aspartyl protease and allows you to edit the protein and set up parameters specific to the protein such as constraints Click on the Protein 1qbt aspartyl protease tab GOLD User Guide Hydrogen Bonding Constraints GOLD features two types of hydrogen bonding constraints A standard hydrogen bond constraint can be used to force a hydrogen bond between a specific protein atom and a specific ligand atom see Standard Hydrogen Bond Constraints A protein hydrogen bond constraint can be used to specify that a particular protein atom should be hydrogen bonded to the ligand but without specifying to which ligand atom see Protein Hydrogen Bond Constraints Standard Hydrogen Bond Constraints A standard hydrogen bond constraint allows a particular ligand atom to be constrained to form a hydrogen bond to a particular protei
193. f the ligand in the input file remember that a given ligand input file may contain more than one ligand This file contains a summary of the fitness scores for all the docking attempts on that ligand The docking attempts are listed in decreasing order of fitness score so the best solution is placed first The file gives total fitness scores and a breakdown of the fitness into its constituent energy terms The example file below corresponds to the fifth ligand in the input file ligand_file mol2 and is therefore called ligand file m5 rnk The solution Mol No 8 corresponds to the file gold_soln ligand file m5 8 mol12 whichis symbolically linked to ranked_ligand_file m5 2 mo12 since it is the second best of the docking attempts for this molecule 127 Fitness list for ligand ligand file molz molecule 5 Mol No Fitness S hb_ext S vdw_ext S hb_int St int 2 46 35 14 08 31 06 0 00 10 44 8 45 10 15 73 29 29 0 00 10 90 9 41 60 12 06 28 45 0 00 9 58 1 39 98 13 61 24 58 0 00 7 43 6 39 05 14 39 27 95 0 00 13 78 4 38 81 10 66 30 47 0 00 13 74 10 36 63 9 08 28 44 0 00 11 56 5 31 65 11 16 22 94 0 00 11 08 7 30 34 4 44 26 59 0 00 10 67 3 28 53 5 83 26 21 0 00 13 34 Average Values 37 80 11 11 27 60 0 00 11 25 e fyou do not wish to save ligand rank files click on Output Options from the list of Global Options given on the left of the GOLD Setup window select the File Format Options tab then disable the Save ligand rank
194. fault button Defining Custom Metal Coordination Geometries It is possible to specify custom metal coordination geometries which can subsequently be used to derive ligand binding points around particular metal atoms GOLD will normalise the size of the custom polyhedron to the appropriate metal chelator distance before matching it to the metal and the coordinating atoms found in the protein Click on the protein tab then select Metals from the list of options given on the left of the GOLD Setup window Click on the Define custom polyhedra button The Define custom metal polyhedra window will appear Define custom metal polyhedra 21x Define Custom Metal Coordination Polyhedra Use this dialogue to specify a custom metal coordination geometry for matching The polyhedron may contain up to nine points The custom geometry may be selected in the metal editor dialogue GOLD will normalise the size of the polyhedron Up to three of these custom polyhedra may be specified Coordinates Point 1 o po boo Point 2 fi fo fo Point 3 A fo gt Point 4 fo fA fo Point ft Point6 f a Point Ly lt Point 8 ft f a Paint 3 Current metal polyhedra Polyhedron 010 100 100 0 10 4 points OK Delete Clear Cancel Custom metal polyhedron may contain up to nine points Each point in the custom polyhedron must be specified using a vector assuming the centre of your polyhedron is at the origin For example t
195. fferent protein file click on the Load Protein button and use the file selection window to choose the protein data file Once selected the chosen protein will be loaded and displayed within the Hermes visualiser e Use the protein tickboxes to determine which proteins are to be docked into e g in the example above all three proteins LACM 1A42 and 1QBT will be docked into e Acceptable protein file formats are PDB and MOL2 3 3 Protonation and Tautomeric States 3 3 1 Adding Hydrogen Atoms to the Protein Using Program Defaults e GOLD uses an all atom model so the protein must have all hydrogen atoms added e To add missing hydrogen atoms to a particular protein select the tab appropriate to the protein you wish to protonate and then select Protonation and Tautomers from the list of available options given on the left of the window e Click on the Add Hydrogens button to protonate the protein e The number of hydrogens added to each atom will be sufficient to satisfy the atom s unfilled valencies 6 GOLD User Guide The hydrogen atom positions will be normalized i e the X H distance will be made equal to the average neutron diffraction value hydrogen atoms are accurately located by neutron diffraction e g C H bond lengths will be set to 1 083 A N H to 1 009 A and O H to 0 983 A It is possible to customise the values for C H N H and O H H normalisation within the Hermes visualiser It is also possible to specify values to n
196. ffold that matches the ligand first will be used This means that it is possible to specify two or more different scaffolds and GOLD will use the scaffold that matches the ligand first This can be useful when docking multiple different series of compounds Setting Up Scaffold Match Constraints To define a scaffold constraint click on Scaffold from the list of Global Options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options The scaffold template file should contain the scaffold molecule or fragment in its docked position i e expressed with respect to the same coordinate frame as the protein and with the coordinates required to place it in the correct pose To specify the scaffold file either enter the path and filename of the file or click on the Scaffold File button and use the file selection window to choose the file The Constraint Weight determines how closely ligand atoms fit onto the scaffold Setting a higher weight will force the ligand to be placed onto the scaffold locations more strictly By default all heavy atoms in the supplied scaffold structure file will be used for matching However it is possible to specify only a subset of those atoms in the scaffold structure these may include non heavy atoms Individual scaffold atoms can be specified by clicking on them in the visualiser Alternatively you can enter the atom numbers as
197. file check box and specify a filename lt filename conf gt By enabling the appropriate check boxes it is also possible to save out the initialised protein structure file see Files Containing the Initialised Protein and Ligand and the cavity atoms file see Defining the Binding Site GOLD User Guide 3 Finish GOLD Configuration 24 x Directory Foaram Files CCDC GOLD Suite GOLD examples tutorial1 ae Save Files IV GOLD conf file gold conf At least one protein has been edited 14CM_protein mol2 IV Protein s I Cavity atoms eavity atoms e If GOLD is run interactively output that is written to the log files are displayed gt Run GOLD 21 x list of ligand logs gold log gold_pratein log gold err Messages ligand log Length of shape fitting list Number of interactions Number of motifs Weight Minimum Include Score Ignore Zero Bits Protein initialisation time total 5 0005 user 5 0004 sys This log file will be updated every 2 seconds Interrupt GA View Solutions Close e A full description of the output files produced by GOLD is available elsewhere see Description of Output Files e The parallel version only gives a summary as it is not possible to track multiple files e If any error conditions are encountered they will be displayed under the gold err tab Note that only fatal errors are reported for the parallel version e You can use the Interru
198. g run see Rescore settings 12 3 Rescore settings e To specify the settings to be used for the rescoring run hit the Rescore Options button This will open the Rescore Options dialog GOLD User Guide 115 i Rescore Options a xi V Perform local optimisation IV Retrieve rotated protein atom positions if available J Use receptor depth scaling Output IV write rescored structures to file IV Replace score tags in file Close e The following rescore options are available Perform local optimisation Enable this check box to minimise the docked ligand pose before rescoring Simplexing is important if you are to obtain meaningful scores Due to the nature of scoring functions one finds that small changes in location or conformation of the pose can have large effects on the calculated score Simplexing can also affect rotatable protein hydrogen atoms see File Containing the Protein Binding Site Geometry Retrieve rotatable H positions from file if available When rescoring a GOLD solution file it is possible to use the optimised positions of the polar protein hydrogen atoms that were generated during the original docking see File Containing the Protein Binding Site Geometry If this option is not switched on or no rotatable H positions are available then the default hydrogen atoms positions specified in the protein input file will be used Use receptor depth scaling This option is only ava
199. gements 142 GOLD was written by Gareth Jones University of Sheffield UK in a DTI LINK collaboration with GlaxoWellcome and the Cambridge Crystallographic Data Centre CCDC Funding was provided by the Biotechnology and Biological Sciences Research Council the Department of Trade and Industry the Medical Research Council GlaxoWellcome Ltd and CCDC Peter Willett University of Sheffield Robert Glen Wellcome Andrew Leach GlaxoWellcome and Jacques Barbanton Lipha Pharmaceuticals are also thanked for significant contributions to the development of GOLD Implementation of the ChemScore Heme Kinase and Astex Statistical Potential scoring functions and the Diverse Solutions code within GOLD is copyright 2001 2007 Astex Therapeutics Ltd We also thank Astex Technology Ltd for their contribution to the water modelling code One of the torsion libraries supplied with GOLD was developed by Gerhard Klebe and Thomas Mietzner BASF GOLD User Guide 20 Appendix A Tutorials In order to familiarise yourself with GOLD it is recommended that you work through the tutorial examples provided Tutorial 1 will go through the process of setting up and running an example docking using the Docking Wizard in some detail subsequent tutorials will be more concise but will introduce other more advanced aspects of the program Tutorial 1 illustrates how to set up protein and ligand files simply using the Hermes visualiser For more compreh
200. he external vdw energy is normally scaled by a factor of 1 375 and summed with the other components to give the total fitness this is to encourage hydrophobic contact between the protein and ligand During a docking run the fitness score may appear to get worse as the docking proceeds This is due to the fact that the effects of poor H bond geometry and close nonbonded contacts are artificially down weighted at early stages of the docking annealing Only the final fitness score i e from the completed docking has any meaning The message Reordering refers to a re ranking of the GA populations caused by the annealing process At the end of the GA run the solution is output and summarised 15 3 Comparison of Docking Solutions GOLD User Guide Following the completion of all docking runs on a ligand the results from the different runs are compared in the ligand log file 131 15 4 132 The file will include a matrix of rms deviations between the various docked ligand positions The rms deviation algorithm takes account of symmetry effects using a graph isomorphism algorithm For example Final Ranking 4 2 5 1 3 RMSD Matrix of RANKED solutions 2 3 4 5 1 4 8 4 7 5 1 10 1 2 4 0 3 1 10 9 3 4 1 10 4 4 11 0 In this case solution number 4 had the largest fitness score this solution will be in gold soln ligand m _4 mo12 which will be symbolically linked to ranked _ligand_m _1 mo12 while solution number 3 had the
201. he distance constraint and whether they belong to the protein or ligand This can be done by clicking on an atom in the visualiser the atom structure type i e protein or ligand will be updated automatically upon selection Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box When specifying atom numbers it is necessary to also set the structure type protein or ligand from the drop down menus The maximum and minimum separation of the constrained atoms must be entered distances are in A and the spring constant must also be specified For example GOLD User Guide 3 GOLD Setup Eoix Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial2 gold conf Load Save Options Protein 1a42 lyase Protonation amp Tautomers Waters Constrain distance from Delete Ligands Flexible Sidechains Protein 7 Atom no 2040 Ligand bd Atom no 29 Soft Potentials Ni Metals Minimum separation 1 5 Constraints gt 5 Distance Maximum separation 3 5 e Spring constant 5 0 jon Protein HBond I Use topologically equivalent atoms Covalent Interaction Motif Distance Protein 2040 Ligand 29 3 5 1 5 5 0 off Help R Run GOLD Run GOLD In The Background Finish Cancel e Ifthe specified ligand atom is topologically equivalent to other atoms in the ligand e g it is one of the oxygen atoms
202. he ligand contains no donors then GOLD can be set up not to dock ligands when the specified constraint is physically impossible to satisfy see Using the Constraint Editor 10 3 3 Setting up Protein H Bond Constraints GOLD User Guide A protein hydrogen bond constraint can be used to specify that a particular protein atom should be hydrogen bonded to the ligand but without specifying to which ligand atom To set up a distance constraint you must first select the appropriate protein tab adjacent to the Global Options tab To define a protein hydrogen bond constraint click on Protein HBond from the list of options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options Specify the protein atoms to be used in the constraint This can be done by clicking on an atom in the visualiser Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box Either a donatable hydrogen atom you must specify the hydrogen atom not the O or N atom to which it is attached or an acceptor can be specified The protein atom should be available for ligand binding i e solvent accessible This constraint does not work with metals The Constraint weight is the strength of bias applied to the formation of a specified hydrogen bond in the least squares mapping algorithm within GOLD The Const
203. he parameters Hydrogen Bonding and FINAL_VIRTUAL_PT_MATCH_MAX are used to set starting and finishing values of max_distance the distance between donor hydrogen and fitting point must be less than max_distance for the bond to count towards the fitness score This allows poor hydrogen bonds to occur at the beginning of a GA run The Van der Waals and Hydrogen Bonding annealing parameters can only be set manually when using user defined GA parameters settings see Using User Defined Genetic Algorithm Parameter Settings Changes to the genetic algorithm parameters should be made with care Click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select GoldScore from the Scoring Function drop down menu The Annealing parameters VdW and H bonding entry boxes can then be used to specify new values Both the vdw and H bond annealing must be gradual and the population allowed plenty of time to adapt to changes in the fitness function 7 3 3 Altering GoldScore Fitness Function Parameters the GoldScore Parameters File A GoldScore parameter file goldscore params is provided in the GOLD_DIR gold directory Parameters can be customised by copying the file editing the copy and instructing GOLD to use the edited file Changes to the scoring function parameters file should be made with care To use a modified goldscore params file click on Fitness and Search Options from the list of Gl
204. hemically sensible This is also done for the covalent link to ensure the geometry around the link is sensible The entry 178 text corresponds to the line number of the angle entry in the BOND_ANGLE_TABLE in the gold params file If you scroll down further you will notice that during the algorithm run the fitness score is broken down into its constituent parts specifically S hb_ext S vdw_ext S hb_int S int In addition to these default scoring terms there is an additional term S cov that is added only when docking covalently Within the BOND_ANGLE_TABLE in the gold params there are also energy terms and the S cov contribution to the score is calculated from this GOLD User Guide This log file will be updated every 2 seconds GOLD User Guide Run GOLD list of ligand logs gold log gold_protein log gold err Messages Doing GA no Operation o 207 328 410 595 1097 Reordering 1125 1318 Reordering 1500 1554 1641 Reordering 1875 2079 z250 2676 3687 3981 Reordering acan Click on the View Solutions button in the Run GOLD window to load the docking results into the 3D view Then hit Close to close the window Fitness 48 49 49 51 55 55 53 56 54 57 59 59 59 63 63 63 64 44 29 49 82 13 20 66 06 30 82 32 25 69 22 68 43 86 S hb_ext S vdw_ext 19 17 15 13 21 26 21 23 22 23 23 23 2
205. hism algorithm is used to determine optimal rms values rms_analysis can be invoked from the command line The structure of the command is dependent on the platform being used Linux lt install_dir gt GOLD_Suite bin rms_analysis method simple complete group_average lt file1 gt mol2 lt file2 gt mol2 lt file3 gt mol2 lt file4 gt mol2 Note This command will only work if users have their GOLD _DIR environment variable correctly set To e g carry out a simple cluster analysis for the files file1 mol2 and file2 mol2 the following command would be used lt install_dir gt GOLD_Suite bin rms_analysis method simple file1 mol2 file2 mol2 Windows via the command prompt lt install_dir gt GOLD gold d_win32 bin rms_analysis_win32 exe method simple complete group_average lt file1 gt mol2 lt file2 gt mol2 lt file3 gt mol2 lt file4 gt molz where lt install_dir gt is the GOLD installation directory If specifying the full path the command will need to be in inverted commas e g C Program Files CCDC GOLD_Suite GOLD gold d_win32 bin rms_analysis_win32 exe method simple complete group_average lt file1 gt mol2 lt file2 gt mol2 lt file3 gt mol2 lt file4 gt mol2 Choose simple for single linkage cluster analysis complete for complete linkage group_average for group average For example the table of rms deviations below for nine dockings of a molecule produces the following cl
206. hm GA parameters see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings GOLD runs for a fixed number of genetic operations crossover migration mutation Therefore reducing the number of GA operations performed during the course of a run will result in GOLD running faster however the search will be less exhaustive e GOLD can decide on the optimal settings to use for a given ligand see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings e To enable automatic GA settings click on the GA Settings option in the list of available options then activate the Automatic radio button The Search efficiency will by default be set to 100 we will use the default settings e We now need to specify an output directory Click on Output Options and specify a directory to which you have write permission This is where the GOLD output files will be written e Now click on the Run GOLD button at the bottom of the interface In the Finish GOLD Configuration window you will be prompted that the GOLD configuration has been updated and needs to be saved click Save to proceed The configuration file name can remain as it is so hit OK to overwrite the existing gold conf This will start 176 GOLD User Guide the GOLD job interactively As the job progresses output will be displayed in the Run GOLD window Any warning messages produced will be displayed under the gold err tab Once the job is complete the
207. hree boxes GOLD User Guide GOLD Setup BE E Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Proteins Define Binding Site Select Ligands Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing E Help R e The approximate radius of the binding site must also be specified If r is the radius Atom select an atom in the visualiser or enter an atom index ZN ZN262 View Point select atoms to define a centroid or edit XYZ Xi f 6 4229 Wi 0 8690 2 15 3360 One or more ligands List of atoms or residues Filename l W View Select all atoms within 10 amp View Reset J Generate a cavity atoms file from the selection Refine Selection IV Detect cavity restrict atom selection to solvent accessible surface J Force all H bond donors acceptors to be treated as solvent accessible Add Definition as a Selection Run GOLD Run GOLD In The Background Finish Cancel the binding site will be defined as all atoms that lie within r of the specified point By default the binding site radius is set to 10 0 This can be changed by entering a value in the box labelled Select all atoms within e Click on the View Selection button to highligh
208. i e the index numbers of the new solutions will continue from the old end point and if solutions were being saved in a single molecule file the solutions of the second set will be appended to that same file e Docking results can also be returned to GoldMine and saved within a new or existing GoldMine DB see Sending Docking Results to GoldMine 5 7 Setting Up Covalently Bound Ligands e GOLD is able to dock covalently bound inhibitors but only if you specify which ligand atom is bonded to which protein atom GOLD supports two types of covalent link A covalent link for use with individual ligands see Setting Up a Single Covalent Link A substructure based covalent link for use with multiple ligands which have a common functional group see Setting Up Substructure Based Covalent Links e Note that it is necessary to use mol2 files when running a covalent docking 44 GOLD User Guide 5 7 1 Method Used for Docking Covalently Bound Ligands GOLD is able to dock covalently bound inhibitors but only if you specify which ligand atom is bonded to which protein atom The program assumes that there is just one atom linking the ligand to the protein e g the O in a serine residue Both protein and ligand files are set up with the link atom included so if the serine O is the link atom it will appear in both the protein and ligand input files Ideally the link atom in both the ligand and the protein will have a free valence
209. ibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Atom Typing A list of Global Options is given on the left of the GOLD Setup window Note that there are a number of setup options that are specific to the protein file thus some options will not be visible until a protein file is read either manually or via a gold conf Click on a configuration option in the list in order to specify the corresponding settings on the right of the GOLD Setup window GOLD User Guide The Hermes visualiser is an integral part of the GOLD interface It is used alongside the GOLD Setup window to prepare input files and for interactive docking setup e g for defining the binding site and the setting of constraints For further information on using the Hermes visualiser refer to the Hermes user guide To simplify the process of setting up a docking a wizard is available which will guide you through the essential configuration steps The wizard can be opened at any stage from the GOLD Setup window by clicking on Global Options on the left of the window then clicking on the Wizard button see Using the GOLD Docking Wizard A number of configuration file templates are also available which contain recommended settings for particular docking protocols see Using Configuration File Templates 2 2 Using the GOLD Docking Wizard GOLD User Guide GOLD has many configuration options To simplify the process of setting up a do
210. ic operator to use next or when to create the starting population of random individuals The random number generator is normally initialised with random seeds it is these seeds that are printed to the go d seed_log file at the end of each docking run The seed file can be used to reproduce identical docking results for repeat runs as long as all other settings are equal To make use of the seed file in this way Copy the required gold seed_log from the original output directory to an alternative location e g the new docking directory Specify the location of the seed file in the gold params file i e open the gold params via the Edit Parameters button in the GOLD interface Find the lines that read Read seeds from SEED FILE if not equal to none Used for debugging SEED FILE none Change the SEED FILE none setting to include the full path to your seed file e g SEED FILE home username new docking dir gold seed_log Then run the docking using the modified gold params file 15 2 Information on the Progress of Docking Runs As each docking run is performed on a ligand the progress of the genetic algorithm is recorded in the ligand log file see Ligand Log File The best most fit individual at any time is listed The total fitness and its component terms are also displayed For GoldScore the internal vdw energy includes the ligand torsional energy T
211. igand Reference File It is possible to supply to GOLD with a file containing a reference ligand e g a crystallographically observed ligand pose The ligand reference file will be used to perform automated RMSd calculations against GOLD solution s For each GOLD solution the resultant RMSd with respect to the reference ligand will be written to the files containing the fitness function rankings i e the ligand rank file rnk and bestranking 1st file Click on Select Ligands from the list of Global Options given on the left of the GOLD Setup window To specify the ligand reference file either enter the path and GOLD User Guide 47 48 filename of the file in the Reference ligand box or click on the button and use the file selection window to choose the file GOLD User Guide 6 Atom and Bond Types 6 1 Atom and Bond Type Overview Each protein and ligand atom must be assigned an atom type which is used for example to determine whether the atom is capable of forming hydrogen bonds GOLD atom typing is based on SYBYL atom types Internally GOLD also uses some additional atom types see Internal GOLD Atom Types SYBYL bond types are also used Correct assignment of atom and bond types is crucial GOLD assigns atom types from the information about element types and bond orders in the input structure file so it is important that these are correct However if for any reason GOLD is unable to deduce an atom type then the
212. igand from its active site activate the tickbox adjacent to A underneath the Extract and Reload header GOLD User Guide 8 GOLD Setup oa J Wizard step 2 Protein setup At this point you have the chance to edit your protein structure if required e g add hydrogens delete waters Global Options 1ACM Protonation amp Tautomers Extract Delete Waters Ligand Extract and Reload Delete Ligands A v Extract Your protein may have one or more ligands occupying the binding site that must be removed before you can peform a docking Extracted ligands are automatically reloaded so they can be used to define the binding site and are written to file for later comparision with docking results Help h lt Back Next gt Cancel Wizard e Now hit the Extract button When prompted save the ligand file e g as ligand mo12 in the folder to which you copied the tutorial1 files The reason for this is that all the files listed in the section Analysis of Output will therefore have the correct name and in all sections below it too e Return to Hermes 3D view you will notice the ligand file has been re loaded separately as A 1ACM underneath the protein entry in the Molecule Explorer e Return to the GOLD Wizard and click on the Global Options tab e Hit Next to proceed to the Define the binding site step GOLD User Guide 147 Defining the Protein Binding Site 148 7 GOLD Setup
213. ilable when rescoring with ChemScore When using receptor depth scaling the score attributed to hydrogen bonds is scaled depending on the depth in the pocket Hydrogen bonds deep in the pocket are rewarded with an increased score while the scores of those closer to the solvent exposed surface are decreased see Receptor Depth Scaling e The following output options are available Write scored structures to file Enable this check box to write out docked ligand solutions after rescoring Solutions will be written to the file rescore mol2 to specify an alternative filename see Rescore Solution File MOL2 or SD output can be specified see Files Containing the Docked Ligand s Solution files will contain the new scoring function terms and can be used with GoldMine If writing of this file is switched off only the rescore log file will be written see Rescore Log File Replace score tags in file When rescoring a GOLD solution file enable this check box to overwrite the list of active residues and the rotated protein hydrogen atom positions generated during the original docking with those resulting from the rescoring run If you select not to replace relevant tags then rescore mo12 will contain both the binding site definition of the original docking and that of the subsequent rescoring run 12 4 Receptor Depth Scaling e In many proteins the cognate ligand forms hydrogen bonds deep in the active site This feature of known binde
214. ile can also be modified see Altering GoldScore Fitness Function Parameters the GoldScore Parameters File The ChemScore fitness function parameters are stored in the chemscore params file which can also be customised see Altering ChemScore Fitness Function Parameters the ChemScore File The Astex Statistical Potential ASP fitness function parameters are stored in the asp params file this can also be customised see Altering ASP Fitness Function Parameters the asp params File 16 4 Customising the Torsion Angle Distribution File 138 It is possible to customise torsion distribution information by copying one of the standard torsion distribution files editing it and instructing GOLD to use the edited file see Editing Torsion Angle Distribution Files GOLD User Guide 17 Context Dependent Help e Context dependent help is available in the GOLD Setup window by clicking the Help button located at the bottom left corner of the window Clicking this button will result in the GOLD user guide being opened on the page appropriate to the task being performed i e the help will reflect the Configuration option currently being set i Help x e Balloon help is also available by clicking on the icon located at the bottom left corner of the GOLD Setup window then clicking on an option within the interface For example selecting and clicking on the protein selection page brings up the following help window gt GOLD Set
215. in 1ase aminotransferase in the example below select Covalent from the list of available options given on the left of the GOLD Setup window and enable the Define covalent docking check box 45 5 7 3 46 3 GOLD Setup Loix Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Protonation amp Tautomers Waters IV Define covalent docking Delete Ligands Se Flexible Sidechains Protein link atom Soft Potentials Metals SEA Ligand link mode Atom C Substructure Interaction Motif Ligand link atom fo Substructure file KEE E J Use topology matching to check test equivalent atoms You can define the covalent link atoms by right clicking in the viewer or by typing in the edit box The ligand link atom can be defined either by a single atom in the ligand or by an atom in a substructure that can be matched against multiple ligands IF using a substructure you must enter the substructure file Help h Run GOLD Run GOLD In The Background Finish Cancel Select Atom as the ligand link mode and define both the Protein link atom and Ligand link atom This can be done by clicking on an atom in the visualiser Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box Setting Up Su
216. ination ne one default or Acceptor A or Coordination TA elucidated Metal M geometries MGD Mg DEF M 4 6 2 05 ZND Zn DEF M 4 5 6 2 09 MND Mn DEF M 4 6 2 06 FED Fe DEF M 4 6 1 98 CAD Ca DEF M 6 7 2 44 COBD Co oh DEF M 6 2 09 GDD Gd DEF M 6 2 44 e The gold params file is stored in lt install_dir gt GOLD Suite GOLD gold go to this directory and open the parameters file using a text editor e The parameters used by GOLD for each metal are listed for explanation of parameters refer to comments in the gold params file Additional metal parameterisation can also be found within the H_BOND TABLE e For our Zn atom GOLD will therefore attempt to match coordination geometries 4 5 and 6 tetrahedral trigonal bipyramidal and octahedral templates onto the GOLD User Guide 159 coordinating atoms found in the protein The template that gives the best match will then be used to generate coordination fitting points e Once you have finished viewing the file close it Manually Specifying Metal Coordination Geometries e Itis possible to manually specify coordination geometries for particular metal atoms see Defining Custom Metal Coordination Geometries This can be useful in allowing non standard metal coordination geometries or to limit the number of possible geometries that GOLD checks i e to overrule the default geometries for the corresponding metal type defined in the go d params file Running GOLD and Analysis of
217. ine work and usually gives comparable predictive accuracy to the slower settings unless the ligand has a large number of rotatable torsions Library screening this sets the search efficiency at 10 This is the fastest setting and as a consequence is the least reliable GOLD User Guide 111 11 3 4 112 e The Minimum number of operations performed during the run will therefore depend on the Search efficiency that is set To ensure that every ligand is subjected to at least a user specified number of operations enable the Min ops check box and specify the minimum number of operations required for every ligand Similarly the maximum number of operations to be carried out can be set manually Using Preset Genetic Algorithm Parameter Settings e The number of genetic operations performed crossover migration mutation is the key parameter in determining how long a GOLD run will take i e this parameter controls the coverage of the search space e When using pre defined GA parameter settings every ligand regardless of it s size and flexibility will be subjected to a specified number of genetic operations e To use a pre defined GA parameter set click on GA Settings from the list of Global Options given on the left of the GOLD Setup window then switch on the button labelled Preset 3 GOLD Setup BE x Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase a
218. inish Cancel e When setting up a distance constraint it is necessary to select both atoms involved in the constraint within the 3D view Alternatively the protein and ligand atom numbers as defined in the MOL2 input files can be typed into the Protein and Ligand windows note you will have to hit return to update the 3D view The maximum and minimum separation of the constrained atoms must also be entered distances are in e During a GOLD run if a constrained distance is found to lie outside the specified bounds a spring energy term is used to reduce the fitness score The spring energy term E kx where x is the difference between the distance and the closest constraint bound and k is a user defined spring constant 174 GOLD User Guide Substructure Based Distance Constraints GOLD User Guide It is possible to apply a distance constraint to multiple ligands which have a common substructure or functional group In order to use a substructure based distance constraint it is first necessary to create a file containing the common substructure in WOL2 format The substructure based constraint forces GOLD to limit the distance between a protein atom and one atom of this functional group During docking the constraint will be applied to any ligands which contain the specified substructure matching is performed on the basis of the atom types and 2D connectivity and the resulting solutions will be biased towards the specified
219. int is incorporated into the least squares fitting routine used by GOLD Thus when least squares fitting is used to dock the ligand by attempting to form hydrogen bonds encoded within the chromosome the constraint is added to the least squares mapping The constraint has a weight of 5 relative to a normal hydrogen bond taken from the chromosome To set up a distance constraint you must first select the appropriate protein tab adjacent to the Global Options tab To define a hydrogen bond constraint click on Hbond from the list of options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options Specify the atoms to be used in the constraint This can be done by clicking on an atom in the visualiser Alternatively you can enter the atom number or PDB sequence number as it appears in the input file directly into the appropriate entry box The hydrogen bond constraint weighting can be altered within the FITNESS FUNCTION section of the GOLD parameters file by changing the value of the parameter CONSTRAINT WT Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor Method Used for Protein H Bond Constraints Protein H bond constraints are applicable to individual protein ligand complexes i e must be set up individually for each protein ligand if performing ensemble docking GOLD wil
220. interface This tabbed window lists all component parts of the visualiser display e g Protein 1fax coagulation factor corresponds to the docked protein and ligands Deselect all radio buttons in the list apart from Protein 1fax coagulation factor and LIM IMA_301_pdbilpg_1 the experimental position of the 1lpg ligand You may wish to hide the H atoms by deactivating the Show Hydrogens tickbox at the top of the Hermes interface if the display is not clear Return to the Docking Solutions tab and select the ligand with the highest fitness score Scores are tabulated under the GoldScore Fitness header in the Molecule Explorer section of the Hermes interface Solutions can be ordered on fitness score by clicking on the GoldScore Fitness header You can scroll through the other docking solutions simply by clicking on them As expected none of the solutions produced in our non flexible run is correct all have the benzyloxy side chain seriously misplaced The top ranked docking has a GoldScore of 63 8592 and is shown below with the true ligand position C atoms coloured grey for reference 187 Running the Flexible Docking and Analysing the Results 188 Clear the display by going to File Close All Files At this point you can choose to load the existing flexible conf to view the results of the flexible docking Alternatively continue with the tutorial to see how to set up a flexible docking Click on the Load button at the top of
221. ion of the protein mol2 input file you will see that the zinc atom is coordinated to three histidine residues 157 Te gli The inhibitor has also been set up in accordance with the guidelines for the preparation of ligands see Essential Steps Open and inspect the file ligand mol2 from the folder to which you copied the tutorial2 files Once you have finished looking at the protein and ligand files close them by selecting File then Close All Files The GOLD Configuration File 158 All of the parameters and settings required to define a particular GOLD job may be saved as a configuration file gold conf see Saving and Reusing Docking Settings This text file will include details of the ligand the protein binding site the fitness function parameter file to be used the torsion distribution file to be used and the genetic algorithm parameters Therefore there is no need to specify protein mol2 and ligand mol2 input files as these will be read in upon opening gold conf A configuration file has been provided for this tutorial The gold conf is loaded by clicking on the main menu option GOLD then Setup and Run a Docking From the resultant pop up window select the Load Existing button you should then browse to the directory containing the tutorial and select the file gold conf from the folder to which you copied the tutorial2 files then hit Open This will automatically load the settings and parameter values for this
222. ions are available by clicking on the More button Automatic enable this to make GOLD automatically calculate an optimal number of operations for a given ligand thereby making the most efficient use of search time e g small ligands containing only one or two rotatable bonds will generally require fewer genetic operations than larger highly flexible ligands see Using Automatic Ligand Dependent Genetic Algorithm Parameter Settings Preset choose from four preset options with varying numbers of operations The larger the number of operations the slower thus more accurate the docking see Using Preset Genetic Algorithm Parameter Settings User defined this option allows you to tailor your GA settings Care should be taken when altering these parameter settings see Using User Defined Genetic Algorithm Parameter Settings and you are recommended to use one of the presets offered Enable automatic GA settings by clicking on the Automatic radio button and ensure the Search efficiency is set to 100 The criteria used by GOLD to determine the optimal GA parameter settings for a given ligand include the number of rotatable bonds in the ligand ligand flexibility i e number of flexible ring corners flippable nitrogens etc the volume of the protein binding site and the number of water molecules considered during docking Details of the exact settings used will be given in the ligand log file gold_ligand_m1 log see Ligand Log Fil
223. ions correspond very closely with the 112i binding mode see below These solutions will have scores of 43 45 The reversed binding mode still appears in some solutions but these invariably have lower scores GOLD User Guide This ends the tutorial GOLD User Guide 197 20 8 Tutorial 8 Generating Diverse Solutions Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial8s toa directory to which you have write permissions The object of this tutorial is to investigate PDB code 3MTH pig hormone complexed with methylparaben insulin The binding site is large and primarily hydrophobic in nature with a small number of acceptor regions The ligand is small thus there is potential for obtaining an incorrect docking pose or poses Consequently GOLD does not perform well when attempting to replicate the binding mode of the ligand 3MTH is an entry in the CCDC Astex validation test set which is available to download as GOLD Validation Sets from http www ccdc cam ac uk SupportandResources Downloads pages ProtectedDow nloadProductList aspx A water molecule which mediates protein ligand binding in the native crystal structure has been reinstated in the protein molecule file all water molecules were removed from the protein files for validation The presence of this water molecule does not improve the standard GOLD docking results significantly i e without using the diverse solutions feature
224. is being run in parallel see Running in Parallel 12 2 Setting Up a Rescoring Run 114 To automatically rescore the results of a docking run with another scoring function you will need to first set up the docking in the normal way Then click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and enable the Rescore check box Select the required scoring function to be used for the rescore from the drop down menu To use a modified scoring function parameters file either enter the path and filename of the Parameters file or click on the button and use the file selection window to choose the file Finally specify the settings to be used for the rescoring run see Rescore settings In the following example ChemScore will be used for the docking and the resulting solutions will be rescored automatically using the Astex Statistical Potential ASP scoring function GOLD User Guide 3 GOLD Setup mE x Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Wizard 4 Templates f I Docking Proteins Define Binding Site Scoring Function GoldScore Select Ligands Waters Parameter file DEFAULT R Ligand Flexibility Fitness amp Search Options Annealing vdw 4 0 H bond las GA Settings Output Options GoldMine rM Rescore Parallel GOLD 5 Constraints Scoring Fu
225. is run through the front end Manually Setting Atom and Bond Types If you do not want to use the automatic atom and bond type assignment available in GOLD you can define the atom and bond types yourself provided that you use MOL2 format This option is useful when you want to set unusual atom types or user defined types GOLD atom typing is based on SYBYL atom types see Appendix B List of Atom and Bond Types SYBYL bond types are also used see Appendix B List of Atom and Bond Types Even if atom types are set manually the automatic atom type assignment software is still run to check the ligand structure for inconsistencies Any errors will be recorded in both the log file and the error file In most cases input types will not be reset If for any reason GOLD is unable to deduce an atom type then the atom in question will be replaced with a dummy atom type Du Bond types must be correctly set see Atom and Bond Type Conventions for Difficult Groups This is normally just a case of checking single and double bonds However the amide bond must be set to the am bond type Also the ar bond type is used for delocalised bonds e g in carboxylate phosphate and guanidinium ions as well as for aromatic bonds Atom types should conform to those expected in SYBYL In particular sp2 oxygen is atom type O 2 sp3 oxygen is O 3 tetrahedral nitrogen is N 3 or N 4 if protonated planar non amide nitrogen is N p 3 and the planar amid
226. itions 113 12 Rescoring 12 1 Overview Different scoring functions may perform better for selected cases You may find for example that ChemScore outperforms GoldScore in ranking actives or one protein class whereas the reverse will apply for other classes Therefore when screening large numbers of compounds rescoring docking poses with alternative scoring functions and considering the best results from each consensus scoring can have a favourable impact on the overall rank ordering of ligands In GOLD it is possible to rescore a single ligand or a set of ligands in one or more files Typically a user will rescore GOLD solution files with an alternative scoring function However it is also possible to score a ligand pose from an alternative source for example from a known crystal structure or a solution from another docking program When rescoring from a source other than a GOLD solution file it will not be possible to use the optimised positions of polar protein hydrogen atoms see Rescore settings Rescoring can be performed automatically after a docking run This will result in the solutions from the docking being automatically scored with another scoring function Alternatively rescoring can be performed independently of the docking e g against an existing set of GOLD solution files or ligand poses from an alternative source see Setting Up a Rescoring Run It is not possible to use the rescore feature if GOLD
227. ive waters and to delete those that are not required see Water Molecules There are not many water molecules co crystallised with the protein and they are not needed for the purposes of this tutorial thus can be deleted Hit the Delete RemainingWaters button When prompted Are you sure you want to delete all the waters hit OK You will be informed that 15 waters have been deleted Extracting and Deleting Ligands GOLD User Guide Before extracting the ligand it is important to ensure the protonation states are correct Hermes makes a best guess at protonation states however as PDB files don t contain atom type information it is not unexpected that Hermes occasionally gets it wrong Minimise the GOLD Setup window and return to the 3D view The most convenient way of editing the ligand structure is to display only the ligand We are going to be docking into chain A so hide all Chains Ligands and Metals apart from Ligand A expand the Ligands tree then deactivate all tickboxes apart from that adjacent to A 145 146 y Hermes EJ Eile Edit Selection Display View Calculate Descriptors GOLD GoldMine Databases Help C Depth Cueing ereo Graphics Objects Show hydrogens V Show unknown atoms Style Wireframe Colour by Element x Picking Mode Pick Atoms oa 1 x x y yt z z x90 x490 y 90 y 90 290 2490 lt gt J T zoom zoom Atom selections kd Molecule Explorer ax Display Movable Descriptors Waters
228. l Potential ASP see Astex Statistical Potential ASP see Astex Statistical Potential ASP see Covalent Docking and Docking with Constraints see Covalent Docking and Docking with Constraints see Protein Protein Clashes see Water Molecules see Internal Energy Offset see Specifying a Ligand Reference File see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP GOLD User Guide Name Gold PLP Chemscore metal Gold PLP Goldscore Hbond Gold PLP DEclash Gold PLP Chemscore protein energ y Gold PLP SBar Gold PLP Chemscore internal corre ction Gold PLP Chemscore covalent Gold PLP constraint Explanation Chemscore Metal binding contribution Goldscore Protein ligand H bond contribution Protein ligand clash penalty to the PLP value Protein energy term to penalise clashes when using flexible sidechains Penalty term for non displacement of active site waters Internal ligand energy offset Covalent bonding contribution to PLP valu
229. l be biased towards finding solutions in which the specified protein atoms form hydrogen bonds The fitness score of a given docking will be penalised by a user specified value c for every protein H bond constraint that is not satisfied i e for every protein atom that you have specified should form a hydrogen bond but does not GOLD assesses the geometry of each required hydrogen bond on a scale of 0 to 1 with 1 denoting perfect If this geometry weight for the constrained Hbond falls below the Minimum H bond geometry weight specified by the user a penalty will be applied to the score for the unfulfilled hydrogen bond i e it will not be considered to be an H bond and will therefore contribute a penalty to the fitness score The magnitude of this penalty is equal to the weight specified for the constraint GOLD User Guide Each trial ligand docking in a genetic algorithm run is generated by a least squares fit of mapping points H bonding or hydrophobic binding points on the protein with complementary points on the ligand The inclusion of a protein H bond constraint will ensure that at least one of the specified protein atoms is included as one of the mapping points i e use of the specified points is enforced at the mapping stage of the algorithm If a ligand simply does not contain sufficient complementary hydrogen bonding atom s to satisfy the specified protein H bond constraints e g you require an H bond to a protein acceptor but t
230. lar regions of the binding site are occupied by specific ligand atoms or types of ligand atom see Region Hydrophobic Constraints Template similarity constraint for biasing the conformation of docked ligands towards a given solution or template see Similarity Constraints Scaffold constraint to place a ligand fragment at an exact specified position in the binding site see Scaffold Match Constraint e To define one of the above constraints ensure the Global Options tab is selected then pick a constraint type from those listed on the left of the GOLD Setup window If individual constraint types are not visible click on the icon next to Constraints to expand the list of options GOLD User Guide 3 GOLD Setup Big ks Conf file C Documents and Settings henderson Desktop ensemble files ensemble conf Load Save Options pdbigpc_full_entry pdbigpd_full_entry pdbigpe_full_entry pdb1qpj_full_entry pdb3ick_a Wizard Templates Proteins Define Binding Site Select Ligands Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options The following constraints are available for definition here GoldMine Parallel GOLD Region reward occupation of specific regions of the binding site ie Constraints e Similarity bias Fragment placement and conformation towards a given solution Similarity Scaffold place exactly a fragment at a specified position in the binding
231. lates the score of hydrogen bonds from the actual distance as opposed to S map where the pre calculated grid points are used The hbond score is the multiplied with the HBOND_ CORRECTION FACTOR making it possible to weight the contribution of hydrogen bonds to the final score this parameter can be changed by editing the asp params file see Altering ASP Fitness Function Parameters the asp params File The S hbond correction is similar to the same term found in ChemScore since the deviation from ideal geometry is taken into account when calculating the score of the hydrogen bond see Hydrogen Bond Terms The S hbond contribution is offset by the score already present in S map GOLD User Guide 71 7 5 5 Covalent Docking and Docking with Constraints e Hydrogen Bond TermsAltering ASP Fitness Function Parameters the asp params FileCovalent docking with ASP is handled by adding a covalent term to the calculated score The implementation is the same as for ChemScore see Covalent Term e Using constraints in conjunction with ASP is carried out using the same principle as with GoldScore and ChemScore see GoldScore 7 5 6 Targeted Scoring Functions e The use of statistical potentials in a scoring function enables the creation of targeted fitness functions to certain proteins This is done by using target specific information when calculating the atom atom potentials Instead of using the information from a general database such as the P
232. ldscore Internal Hbond Gold Goldscore Internal Vdw Gold Goldscore Internal Torsion Gold Goldscore Covalent Energy Gold Goldscore Constraint Score Gold Goldscore Internal Correction Gold Goldscore Protein Energy Gold Goldscore SBar Gold Goldscore Reference RMSD Chemscore Gold Chemscore ZeroCoef Gold Chemscore Rot Gold Chemscore Fitness Gold Chemscore Hbond 222 Explanation When docking into protein ensembles this is a numerical identifier given to each initialised protein The Gold Ensemble ID corresponds to the number in the output protein file i e gold protein 1 mol2 gold protein 2 mol2 Total GoldScore fitness value of docked ligand Protein ligand H bond contribution to GoldScore value Protein ligand vdw contribution to GoldScore value Internal ligand intramolecular H bond contribution to GoldScore value Internal ligand vdw contribution to GoldScore value Internal ligand torsion strain contribution to GoldScore value Covalent bonding contribution to Goldscore value Constraint contribution to GoldScore value Internal ligand energy offset Protein energy term to penalise clashes when using flexible sidechains Penalty term for non displacement of active site waters RMSd of solution against reference ligand The Chemscore zero coefficient Rotatable bond freezing term contribution to Chemscore value Total Chemscore fitness value of docked ligand Protein liga
233. lex with a number of cyclic urea inhibitors have been determined It has been observed that the central urea moiety is anchored in the active site of the protease by six key hydrogen bonds Two hydrogen bonds between the urea oxygen atom and the protein backbone peptide groups of Ile50 and Ile50 shown below Four hydrogen bonds between the cyclic urea diol and the carboxylates of the catalytic aspartate of the protein residues ASP25 shown below e Protein H bond constraints can be used in order to attempt to reproduce these key interactions during docking e Specify that either oxygen atom of the carboxylate group of Asp25 in chain A should form a hydrogen bond to the ligand by clicking on one O atom then the other Note If you have problems locating Asp25 in chain A you can expand the protein tree in the Molecule Explorer i e by clicking on the adjacent to the words Protein 1abt aspartyl protease Expanding the Chains tree then A will give a breakdown of all residues in chain A You can then right click on Asp25 to modify the display settings and make it stand out e Once both carboxylate atoms have been selected they will be highlighted with cyan spheres GOLD User Guide 167 e Inthe constraints window the default settings for Constraint weight and Minimum H bond geometry weight are given 10 0 and 0 005 respectively Select Add to accept these values The specified constraint will appear in the Constrain
234. lick on the corresponding motif number column then hit the Delete Motif button The maximum number of motifs that can be defined is 20 In the example below 3 motifs have been defined Motif M1 features Glu810 as an acceptor Leu83N as a donor Leu830 as an acceptor but does not include Leu830 as a CHO acceptor Motif M2 features Glu810 as an acceptor Leu83N as a donor and Leu830 as a CHO acceptor Motif M3 features Glu810 as an acceptor and Leu83N as a donor The lipophilic interactions are included in all the motifs and are expressed as the frequency of that interaction as observed in the set of complexes originally used to identify the motifs 105 106 GOLD Setup Big ks Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial2 gald conf Load Save Options Protein 1a42 lyase Protonation amp Tautomers Waters Add Interaction Add Motif Delete Ligands Delete Interaction Delete Motif Flexible Sidechains Soft Potentials Metals Donor Lipohilic Acceptor Lipohilic Donor Acceptor CHO Lipohilic Donor 10 Lipohilic Residue Chain Name Atom 7 4 TYR Constraints Define up to 10 interactions and 20 motifs Covalent Interaction Motif Type OH Mi M2 Reset View Atoms M3 1 65 4 ALA CB 1 62 4 ASN 1 20 PHE 198 LEU 198 LEU 198 LEU 141 LEU 121 91
235. ligands like methotrexate curl up 8 5 Flipping Planar Nitrogens e Click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Flip all planar R NR1R2 check box to allow planar trigonal nitrogens in the ligand bound to sp2 carbons to flip between cis and trans conformations during docking otherwise they will be held fixed at the input geometry e Itis possible to independently control the behaviour of both ring NHR and ring NR1R2 groups during docking The following options are available for each Flip allows ring NHR and ring NR1R2 to flip i e rotate 180 deg during docking Rotate use this option to allow free rotation of ring NHR or ring NR1R2 groups during docking Fix This fixes ring NHR or ring NR1R2 bonds at their input conformation 8 6 Protonated Carboxylic Acids e Click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Flip protonated carboxylic acids check box e Protonated carboxylic acids can then either be allowed to flip i e rotate 180 deg or rotate freely during docking e ifthe Flip protonated carboxylic acids check box is not switched on then these groups will be held rigid at their input conformation 8 7 Using Torsion Angle Distributions 8 7 1 Enabling Use of Torsion Angle Distributions e Torsion angle distributions extracted from the Cambridge Structural Database
236. ll contain a Loading protein section for each initialised protein i e each protein in the ensemble and there will be an Active Molecule Initialisation section for all initialised waters molecules in the ensemble GOLD User Guide Molecule loaded from file C 31ek 3LCK_HOH1015 mo12 Molecule name 3LCK_HOH1015 Ld Protein scores for each ligand are contained with the ligand log output file You can see below that for this particular ligand protein 1 scores highest Fitness S hb_ext S vdw_ext S hb_int Siint intcor protein 1 101 72 33 83 55 46 0 00 8 37 5 10 protein 2 100 97 33 71 55 28 0 00 8 76 5 10 protein 3 88 70 25 33 48 49 0 00 3 30 5 10 4 3 5 Caveats of Docking into Ensembles Although it is possible to specify rotatable side chains when docking an ensemble these sort of movements can be captured in an additional protein mode that can be added to the ensemble This might be worth considering before setting any side chains as flexible Each protein is assigned an index number by GOLD when the ensemble docking is carried out It is possible to rescore an ensemble docking however if a separate conf file is used from the original docking it is essential that the order the proteins are listed in is maintained in the rescore run If the protein order is not retained the rescore will not run In ensemble docking it is possible to define constraints on individual protein models or on all protein models Constraints work by
237. ll vary depending on how many proteins are loaded 3 GOLD Setup Bi ks Wizard step 1 Select one or more proteins Either choose a protein already loaded in the visualiser or load a new file Options 1acm Wizard steps N X 5 5 1 Select a protein Select proteins to use Load Protein Superimpose Proteins 2 Protein setup 3 Define the binding site 4 Configuration template 5 Select ligands 6 Choose a fitness function 7 GA search options 8 Finish 1acM Protein score offset ensemble docking only Score Offset Help h lt Back Next gt Cancel Wizard e The wizard will guide you through the steps required to configure a docking At each step follow the instruction provided Once a step has been completed click on the Next button to proceed to the next configuration step or Back to return to the previous step To cancel the wizard click on the Cancel Wizard button e Tutorial 1 describes in detail how to use the GOLD wizard see Tutorial 1 A Step By Step Guide to Using GOLD GOLD User Guide 3 Setting Up the Protein s 3 1 Essential Steps Protein setup is the same whether an individual protein or an ensemble of proteins is being used You can either input the whole protein structure to GOLD or just those residues that are in the active site region The latter leads to somewhat shorter run times since both protein initialisation and cavity detection will be quicker If you i
238. lling program to create and edit starting models Commonly used molecular modelling environments include SYBYL http www certara com products molmod sybyl x Insight Il or Cerius2 http www accelrys com Discovery Studio http accelrys com MOE http www chemcomp com Predicting how a small molecule will bind to a protein is difficult and no program can guarantee success The next best thing is to measure as accurately as possible the reliability of the program i e the chance that it will make a successful prediction in a given instance For that reason GOLD has been tested on a large number of complexes extracted from the Protein Data Bank The overall conclusion of these tests was that the top ranked GOLD solution was correct in 70 82 of cases GOLD offers a choice of scoring functions GoldScore see GoldScore ChemScore see ChemScore ASP see Astex Statistical Potential ASP CHEMPLP see Piecewise Linear Potential CHEMPLP and User Defined Score which allows users to modify an existing function or implement their own scoring function see User Defined Scoring Function Different values of the genetic algorithm parameters may be used to control the balance between the speed of GOLD and the reliability of its predictions see Balancing Docking Accuracy and Speed GOLD will only produce reliable results if it is used properly and correct atom typing for both protein and ligand is particularly important se
239. lt all protein atoms within 5 0 of each selected ligand are used for the binding site definition This can be changed by entering a new value in the box labelled Select all atoms within e Residues that have at least one of their atoms included in the binding site definition will be highlighted in the Hermes visualiser When entering a new value in the Select all atoms within box it is necessary to hit the enter key before the visualiser will update to reflect the changes made e After visual inspection you may wish to manually refine the binding site definition To do this switch on the check box labelled Generate a cavity atoms file from the selection By enabling this option the binding site definition will automatically be expanded to include all atoms in the existing definition plus all the atoms of their associated residues To manually refine this selection click on the Refine Selection button to open the Refine Binding Site Selection dialogue All residues included in the binding site definition are listed Residues can then be added or removed from the selection by clicking on atoms in the Hermes visualiser 16 GOLD User Guide e The cavity atom selection can be saved as a protein atom subset and viewed within Hermes To do this click on the Add Definition as a Selection button You can then highlight the atoms belonging to the subset by picking the required subset from the Atom Selections pull down menu which is situated above the vi
240. lts into Hermes by clicking on the View Solutions button in the Run GOLD window We no longer need the Run GOLD window so close it by hitting the Close button e The docking results will be loaded into the Hermes visualiser The different explorer windows in Hermes are dockable so move any windows that may obscure your view of the docking results GOLD User Guide 161 162 The protein file now contains dummy atoms connected to the metal which represent idealised metal coordination positions These can be visualised by activating the Show unknown atoms tick box at the top of the Hermes window At locations where GOLD is missing a coordination site i e coordination points not bound to the protein virtual coordination points are added These coordination points are then used as fitting points that can bind to acceptors Inspect how well the docked inhibitor fits within the protein binding site The docking solutions are listed in the Docking Solutions tab of the Molecule Explorer their CHEMPLP fitness score is listed alongside Click on the top solution and holding the left mouse button down drag with your mouse to the final solution in the list selecting all solutions The ligands all coordinate in the same way to the metal i e via the sulfonamide moiety The zinc shown in blue is coordinated to the protein via three histidine residues In the example shown below the remaining zinc coordination site is used to bind the inhibitor
241. lve the metal binding to for example carboxylate ions deprotonated histidines i e negatively charged and phenolates Therefore metals can be considered to bind to H bond acceptors and the metal will compete with H bond donors for interaction Consequently GOLD uses the following approach for handling metals Virtual coordination points are added at locations where GOLD is missing a coordination site These coordination points are then used as fitting points that can bind to acceptors 3 9 6 Heme Containing Proteins 24 The paper Kirton et al Proteins Structure Function and Bioinformatics 58 836 844 2005 describes the use of ligand specific iron parameters in the context of docking to heme containing proteins This extended metal parameterisation is available for the fine tuning of metal interactions so that e g metal ligand interactions can specifically be addressed depending on the metal contact The protein does not need to be set up in a special way to make use of these parameters however the standard set up should be followed see Preparing a Protein Input File which Contains a Metal lon Further information on setting up a GOLD run with these settings is available see Heme Scoring Function GOLD User Guide 4 Protein Flexibility Protein flexibility can be handled in one of three ways in GOLD by allowing side chains to rotate within user defined bounds during docking see Side Chain Flexibility b
242. mScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File GOLD User Guide 65 Term rq Rigel Rmax O lipo 7 4 5 66 interaction is deemed to occur Lipophilic parameters in ChemScore Meaning Name in ChemScore File Default Value The actual distance between the pair of Calculated for each atom lipophilic atoms in A atom pair The ideal atom atom distance separation LIPO RI 4 1 The maximum separation beyond which no LIPO_R2 7 1 The Gaussian smearing sigma associated with LIPO R_ SIGMA 0 1 this term The difference between the metal and lipophilic parameterisation is that the lipophilic term is scored over a much longer range Lipophilic atoms are defined as non accepting sulphurs non polar carbon atoms polar carbon atoms are carbon atoms attached to two or more polar atoms and non ionic chlorine bromine and iodine atoms The lipophilic term has a regression coefficient associated with it v3 By default this is set to 0 117 The name of this coefficient in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File is LIPO COEFFICIENT Rotatable Bond Freezing Term The following formula is used to estimate the entropic loss that occurs when single acyclic bonds in the ligand become non rotatable upon binding 1 MES Ta 4 Pit N rot 2 F EAFF VD N is the number of frozen rotatable bonds in the
243. message Finished Docking Ligand ligand mol2 will appear in the gold_ligand_m1 log tabbed view Once you view this message load the results into Hermes using the View Solutions button then close the Run GOLD window using the Close button Analysis of Output GOLD User Guide A file called bestranking I st is written to the specified output directory for batch jobs Open this file and inspect it using a text editor it gives a continuous summary of the best solution that has been obtained for each docked ligand The listed file names correspond to the names of the files containing the best solution found for each ligand The file gives total fitness scores and a breakdown of the fitness into its constituent energy terms An additional constraint scoring term DE con is also listed For docking solutions which satisfy the specified distance constraint the contribution from this scoring term will be 0 00 However for solutions in which the constrained distances lie outside the specified bounds a negative DE con score will be applied thus reducing the overall fitness Further details relating to substructure based constraints are given within individual ligand log files Your output directory should contain ten ligand log files gold_ligands_m log one for each ligand Open and inspect the ligand log file corresponding to the first ligand in the input file i e gold_ligands_m1 log This file will contain the distance bounds as specified in
244. minotransferase Wizard Templates Automatic Proteins Define Binding Site Preset Select Ligands Waters User defined Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing E 100 000 operations 50 000 30 000 10 000 Help R Run GOLD Run GOLD In The Background Finish Cancel e Select the required number of genetic operations from those listed e 100 000 operations deliver high predictive accuracy but are relatively slow These settings are recommended for use with large highly flexible ligands or for research applications where speed of docking is not an issue and optimal accuracy is required e 50 000 operations and 30 000 operations are progressively quicker predictive reliability will fall off but quite slowly These setting are recommended for use with GOLD User Guide compounds containing up to six flexible bonds and or ring corners see Ring Conformations 10 000 operations will give comparable predictive accuracy to the slow 100 000 operations setting when docking small rigid ligands These settings are recommended for use with ligands containing one or two rotatable torsions and for virtual screening work 11 3 5 Using User Defined Genetic Algorithm Parameter Settings Individual GA parameters can be specified manually However it is recommended that you use the automatic liga
245. ms e g it is one of the oxygen atoms of an ionised carboxylate group GOLD will automatically compute the constraint term using whichever of the equivalent atoms gives the best value Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor 10 3 Hydrogen Bond Constraints GOLD User Guide Two types of hydrogen bond constraints may be specified A hydrogen bond constraint see Setting Up Hydrogen Bond Constraints which can be used to force a hydrogen bond between a particular protein atom and a particular ligand atom A protein hydrogen bond constraint see Setting up Protein H Bond Constraints which can be used to specify that a particular protein atom 95 10 3 1 10 3 2 96 should be hydrogen bonded to the ligand but without specifying to which ligand atom Setting Up Hydrogen Bond Constraints Hydrogen bond constraints are applicable to individual protein ligand complexes i e must be set up individually for each protein ligand if performing ensemble docking A ligand atom may be constrained to form a hydrogen bond to a particular protein atom One atom should be a donatable hydrogen atom you must specify the hydrogen atom not the O or N atom to which it is attached and the other should be an acceptor The protein atom should be available for ligand binding i e solvent accessible Note that this constraint does not work with metals The constra
246. ms representing idealised coordination positions These dummy atoms will be connected to the metal ion Any unoccupied coordination points will then be available for ligand binding see Metal Ligand Interactions GOLD User Guide 21 3 9 3 22 Specifying Metal Coordination Geometries Manually It is possible to manually specify coordination geometries for particular metal atoms This can be used to allow non standard metal coordination geometries or to limit the number of possible geometries that GOLD checks i e it is possible to overrule the default geometries for the corresponding metal type defined in the gold params file see Automatic Determination of Metal Coordination Geometries Metal ions are protein specific so first activate the relevant protein tab adjacent to the Global Options tab e g Protein 1a42 lyase in the example below Click on Metals from the list of options given on the left of the GOLD Setup window gt GOLD Setup Bee Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gald conf Load Save Options Protein 1a42 lyase Protonation amp Tautomers Waters Coordination geometries atom S J Tetrahedral Delete Ligands Hg 2039 Default Zn 2040 Default I Trigonal bipyramid Flexible Sidechains Soft Potentials I Octahedral Constraints Covalent J Capped trigonal prism Interaction Motif J Square prism J Icosahedral J Dodecahedral J Custom geomet
247. n atom Click on the triangle next to Contraints in the Protein 1qbt aspartyl protease tab this will expand the tree to list all constraint options available Select HBond from the list of constraint types When specifying a hydrogen bond constraint the ligand and protein atoms involved in the constraint need to be selected in the 3D view Clicking on the atoms simultaneously selects them selected atoms will be surrounded by a cyan sphere and enters the relevant atom IDs into the constraints dialogue One of the atoms must be an H bond donor and the other should be an acceptor The protein atom must also be available for ligand binding i e solvent accessible Once defined an H bond constraint is incorporated into the least squares fitting routine used by GOLD to dock the ligand The constraint has a weight of 5 relative to a normal hydrogen bond Thus the docking will be biased towards solutions which include the specified hydrogen bond The hydrogen bond constraint weighting can be altered within the Fitness Function section of the GOLD parameters file by changing the value of the parameter CONSTRAINT WT Protein Hydrogen Bond Constraints A protein hydrogen bond constraint can be used to specify that a particular protein atom should be hydrogen bonded to the ligand but without specifying to which ligand atom see Setting up Protein H Bond Constraints General Methodology GOLD User Guide Click on the triangle next to Con
248. n below GOLD User Guide 27 28 Edit Rotamer Library TYR99 _ O x TYR99 Chil Chi2 0 0 90 90 90 90 180 180 106 j J51 Reset Reset m Rotamer Library Operations Library Rigid Free Crystal 2 From renee I Improper Delta Chi1 is the first rotatable torsion in the side chain In this example it corresponds to rotation around Ca CB so the atoms will be the backbone N atom 1286 CA 1287 CB 1290 and CG 1291 Chi2 is the second rotatable torsion and corresponds to rotation around CB Cy so the atoms are CA 1287 CB 1290 CG 1291 and CD1 1293 Thus rotamer1 specifies the first set of allowed values for chi1 and chi2 In this example this is chil 60 chi2 90 Rotamer2 specifies the second set of allowed values In this example de ta1 10 and specifies the allowed range deltal chi1 to delta1 chi1 while delta2 10 15 and specifies the range chi2 10 to chi2 15 In summary the effect of these two rotamers is therefore to allow Tyr99 to adopt the conformation of precisely chil 60 chi2 90 or any conformation in the range chil 55 to 75 chi2 100 to 70 Each rotamer therefore describes one allowed conformation of the side chain as defined by the torsion angles values chi1 chi2 etc and their allowed ranges delta1 delta2 etc Rotamers can be defined in the following ways Setting a side chain to be rigid To fix
249. n is selected e Anumber of additional options are available by clicking on the More gt gt button By default the Allow early termination check box should be switched on Remove the tick next to Allow early termination This will switch off early termination to ensure as many solutions as possible are explored 214 GOLD User Guide 98 GOLD Setup Ee Wizard step 6 Choose a fitness function GOLD offers several different scoring functions the original GoldScore is the default Please choose the one you wish to use for this docking Global Options 1TBF 1t9s 1xpd 2chm Wizard steps Docking 1 Select a protein 2 Protein setup Scoring Function CHEMPLP Ne 3 Define the binding site 4 Configuration template Parameter file DEFAULT 5 Select ligands 6 Choose a fitness function 7 GA search options 8 Finish Less lt lt C Rescore Scoring Function CHEMPLP C Allow early termination Early Termination Options Generate diverse solutions Diverse Solution Options Use the internal ligand energy offset Read hydrophobic fitting points File fit_pts mol2 Vie GOLD parameter file DEFAULT Edit Help h lt Back Next gt Cancel Wizard e Hit the Next button to proceed to the Genetic Algorithm search options dialogue Specifying GA Settings e GOLD optimises the fitness score using a genetic algorithm GA A number of parameters control the precise operation of the genetic algorithm The s
250. n parameters file located within the GOLD_DIR gold directory To employ this files click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select ChemScore from the Scoring Function drop down menu Then either enter the path and filename of the Scoring function parameter file or click on the button and use the file selection window to choose the chemscore kinase params file This will enable the recognition of activated CH groups for hydrogen bonding Active CH groups are those in aromatic rings next to nitrogens e g the CHs in an imidazole ring These groups are recognised both in the ligand and protein active site For further details please refer to Virtual Screening Using Protein Ligand Docking Avoiding Artificial Enrichment see References 7 8 2 Heme Scoring Function GOLD User Guide The heme scoring function is available for both GoldScore see GoldScore and ChemScore see ChemScore By default GOLD makes no distinction between different H bond acceptors in terms of their strength of interaction with the metal A recent publication by Kirton et al S B Kirton C W Murray M L Verdonk and R D Taylor Proteins Structure Function and Bioinformatics 58 836 844 2005 demonstrated how metal parameters can be set up in GOLD for both GoldScore and ChemScore to take account of different H bond acceptor types Kirton et al described the use of ligand specifi
251. n the unfulfilled valency at the substitution point which must not be blocked by hydrogen Unlike the template similarity constraint which will bias the docking by adding an energy term to the score based on the similarity between the ligand being docked and the template provided this constraint is enforced at the mapping stage in GOLD Ligand placements are generated using a best least squares fit with the scaffold heavy atom positions i e this constraint forces all atoms on the matching portion of the ligand to lie very close or coincident with the corresponding scaffold There is no S con contribution to the fitness score to bias dockings How closely ligand atoms fit onto the scaffold is governed by a user specified weight Setting a higher weight will force the ligand to be placed onto the scaffold locations more strictly A default weight of 5 0 is used Setting a high weight can have a detrimental effect on the fitness score if the placement results in e g bad protein ligand clashes If desired values below 1 can be used to achieve a more lenient overlay Symmetry effects such as the flipping of a phenyl ring by 180 degrees are not taken into account during matching of the ligand onto the scaffold Therefore a scaffold that will give a unique match should ideally be provided For a given ligand it is not possible to match multiple scaffolds at the same time Scaffolds are evaluated in the order supplied by the user and the sca
252. n the Edit button The resulting Edit Rotamer Library dialogue should then be used to set the rotational parameters for the selected side chain see Defining Rotamers Alternatively select a side chain within Hermes by right clicking on it and selecting Set flexibility parameters from the drop down list The resulting Edit Rotamer Library dialogue should then be used to set the rotational parameters for the selected side chain see Defining Rotamers e A maximum of 10 flexible side chain can be defined e Once rotational parameters have been specified the Status of those side chains made flexible will be updated in the list To highlight in the Hermes visualiser only those side chains that have been made flexible click on the Highlight flexible button To highlight all side chains in the defined binding site click on Highlight all and to remove all highlighting click on the Highlight none button GOLD User Guide 4 1 3 Defining Rotamers e Once a side chain has been specified as flexible you will be required to define one or more allowed rotamers Each rotamer specifies the torsion angles that are permitted to vary and the allowed values or ranges of values for those torsion angles Up to 50 rotamers can be defined for each flexible side chain e Rotamers are defined using the Edit Rotamer Library dialogue which is opened when selecting a side chain see Specifying Flexible Side Chains For example consider the side chain TYR99 show
253. n the Run GOLD window If you have already closed the Run GOLD window this file can be found in the output directory specified see The GOLD Configuration File and can be read using a text editor e The gold_protein log file contains details of the parameterisation of the protein and the determination of the ligand binding site Information relating to the metal and the determination of the coordination geometry will also be given Run GOLD list of ligand logs gold log gold_protein log gold err Messages ligand log 153 2162 2164 Number of active atoms 143 Metal coordination spheres z initialised Processing metal ion no z 2040 mame ZN type Zn Coordinating residues HISS4 atom 732 HIS96 atom 753 HIS119 atom 930 tetrahedral RMSd 0 299 trigonal _bipyramid RMSd 0 233 octahedral RMSd 0 431 Matched coordination geometry z trigonal_bipyramid RMSd of matched geometry 0 233 Protein donor atoms S ah 711 729 1542 750 479 1549 40 517 1537 1544 19 Protein acceptor atoms 9 s 920 5232 737 1547 514 1549 An 1554 492 This log file will be updated every 2 seconds Interrupt GA View Solutions Close e You will see that GOLD has matched a trigonal bipyramid geometry as this geometry has the lowest RMSD e Further information about the contents of the go d_protein log file is given elsewhere see Protein Log File Files Containing the Protein and Docked Ligands e Load the docking resu
254. nalisation of near optimal conformations Performing a few cycles of molecular mechanics minimisation before docking may help to take the ligand close to its local potential energy minimum GOLD User Guide 85 9 Ligand Search Options 9 1 Internal Energy Offset Click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window The Use the internal ligand energy offset check box is switched on by default Enabling this option results in the internal energy terms internal torsion internal vdw and internal Hbond being corrected according to the best energy encountered for these terms during the run By applying this correction the internal energy will be calculated with respect to that of a close to optimal non bound structure thereby taking into account any irreducible internal energy For each scoring function the ligand energy correction value is written to the docked solution files in the tag lt Gold lt scoring function gt Internal Correction gt This is the best i e minimum energy value encountered For all scoring functions the best value encountered is subtracted from the ligand score or energy value before being passed to the final energy term The rnk file is corrected at the end of a run with the best energy encountered after all docking attempts on a particular ligand individual solution files are not Therefore you may observe small deviations for the best energy fo
255. nction z Atom Typing Parameter file DEFAULT eS Rescore Options IV Allow early termination Early Termination Options J Generate diverse solutions Diverse Solution Options I Use the internal ligand energy offset J Read hydrophobic fitting points File Fit _pts mol2 vA View GOLD parameter file DEFAULT oe Edit Help R Run GOLD Run GOLD In The Background Finish Cancel e To rescore an existing set of GOLD solution files or ligand poses from an alternative source i e without first running a docking enable the Rescore check box only and select the required scoring function to be used for the rescore from the drop down menu To use a modified scoring function parameters file either enter the path and filename of the Parameters file or click on the button and use the file selection window to choose the file e Rescoring in this way requires essentially the same information as a normal docking run You will therefore need to Provide a prepared protein input file see Specifying the Protein File or Files Define the binding site preferably the same definition that was used for the original docking see Defining the Binding Site Specify the ligand s you wish to rescore see Specifying the Ligand File s Specify the fitness function to be used for the rescoring see Selecting a Fitness Function e Finally specify the settings to be used for the rescorin
256. nd H bond contribution to Chemscore value See see Interpreting Ensemble Docking Output see GoldScore see GoldScore see GoldScore see GoldScore see GoldScore see GoldScore see GoldScore see GoldScore see Internal Energy Offset see Protein Protein Clashes see Water Molecules see Specifying a Ligand Reference File see Overview see Rotatable Bond Freezing Term see Overview see Hydrogen Bond Terms GOLD User Guide Name Gold Chemscore Lipo Gold Chemscore Metal Gold Chemscore internal_Hbond Gold Chemscore DEClash Gold Chemscore DEInternal Gold Chemscore DG Gold Chemscore Covalent Gold Chemscore Constraint Gold Chemscore CHOScore Gold Chemscore I nternal Correctio n Gold Chemscore Protein Energy Gold Chemscore SBar Gold Chemscore Reference RMSD Astex Statistical Potential ASP Gold ASP Fitness Gold ASP ASP Gold ASP Map Gold ASP Hbond Gold ASP Metal GOLD User Guide Explanation Protein ligand lipophilic contribution to the Chemscore value Metal binding contribution to Chemscore value Internal ligand intramolecular H bond contribution to Chemscore value Protein ligand clash penalty to the Chemscore value Internal ligand torsional strain penalty to the Chemscore value Free energy change that occurs on ligand binding contribution to Chemscore value Covalent bonding contribution to Chemscore value
257. nd dependent settings see Using Automatic Ligand Dependent Genetic Algorithm Parameter Settings or one of the pre defined GA parameter sets see Using Preset Genetic Algorithm Parameter Settings as opposed to altering individual parameters because the optimum values of the parameters are highly correlated To manually specify individual GA parameter values click on GA Settings from the list of Global Options given on the left of the GOLD Setup window then switch on the button labelled User defined 3 GOLD Setup Mie x Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Wizard Templates C Automatic Proteins Define Binding Site Preset Select Ligands Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing User defined Population size 100 Selection pressure fa Number of operations ooo tt Number of islands f Niche size jz i Crossover Frequency jo Mutation Frequency js Migration frequency fo Ft Help R Run GOLD Run GOLD In The Background Finish Cancel GOLD User Guide The values for individual GA parameters can be specified using the appropriate entry box A definition of the individual genetic algorithm parameters are provided in see Appendix D Genetic Algorithm Parameter Defin
258. nd instructing GOLD to use the edited file To use a modified gold params file click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window Then either enter the path and filename of the GOLD parameter file or click on the button and use the file selection window to choose the file If the parameter file is set to DEFAULT then the standard GOLD distribution parameter file is copied to the current directory GOLD gets the location of the parameter file from the configuration file line param file lt parameter file location gt This is most easily defined using the Parameter File button in the front end GOLD User Guide The format of the parameter file is quite strict incorrect editing may cause GOLD to behave in unexpected ways or even to crash Because of the large number of parameters no guarantee can be given that the program will behave reliably with anything other than the default parameterisation For more information see the comments in the parameter file gold params 7 8 Targeted Scoring Functions 7 8 1 Kinase Scoring Function Weak CHO interactions can be accounted for by inclusion of a ChemScore term that calculates a contribution for weak hydrogen bonds This term can be useful when dealing with particular proteins e g most kinases contain weak N heterocycle CH 0 hydrogen bonds This term can be enabled by using the chemscore kinase params scoring functio
259. ned distrbutions The file to be used is specified in the Ligand Flexibility window under Global Options in Advanced options During the Docking e As the job progresses output will be displayed in a Run GOLD window e This is a tabbed view that allows inspection of several files list of ligand logs gold log gold_protein log gold err Messages and ligand log e Any error or warning messages produced will be displayed under the gold err tab This may contain a number of warning messages relating to the GOLD atom type assigner These messages can be safely ignored e Once the job is complete the docking results can be loaded into Hermes by clicking on the View Solutions button in the Run GOLD window Keep the Run GOLD window open GOLD User Guide 153 Analysis of Output In addition to the files that can be inspected in the Run GOLD window the specified output directory see Specifying Ligand Solution File Formats and Directories will contain a number of files including Files containing the initialised protein and ligand gold_protein mol2 and gold_ligand mol 2 Files containing the docked ligand gold_soln_ligand_m1_n mol2 Files containing fitness function rankings igand_m1 rnk and bestranking Ist Protein and ligand log files gold_protein log and gold_ligand_m1 log Files containing error messages gold err this file will be empty if no errors are found Some of these output files will be dealt with in de
260. nfiguration dialogue Run the Docking 216 Before hitting the Run GOLD button click on the Advanced button on the bottom right of the interface This takes us to the standard GOLD set up interface Select Output Options under Global Options Click on the button next to Output directory and specify a directory to which you have write permission this is where the GOLD output files will be written GOLD User Guide 8 GOLD Setup a Conf file Load Save Global Options 1TBF 1t9s 1xp0 2chm Wizard File Format Options Information in File Selecting Solutions Templates Proteins Output file format Same as input SD file Mol2 Define Binding Site re P Select Ligands Output directory ensemble_docking output Configure Waters Create output sub directories for each ligand Ligand Flexibility Fitness amp Search Options F Save ligand rank rnk files GA Settings Output Options V Save ligand log files GoldMine v Save initialised ligand files Parallel GOLD Cak Save solutions to one file Atom Typing Use alternative bestranking lst filename Create links for different binding modes based on RMSD clustering Distance between dusters 0 75 A Help R RunGOLD Run GOLD In The Background Finish Cancel e We have now finished setting up our docking so click on the Run GOLD button at the bottom of the GOLD interface You will be presen
261. ng With No Soft Potential Applied GOLD User Guide The files 1 2i_prot mol2 and 1x7r_prot mol2 are the protein models derived from the pdb entries 112i and 1x7r respectively 1 2i lig mol2 is the ligand structure obtained from and in the same frame of reference as 112i The GOLD configuration file gold_1 2i_1I2i conf is set up to dock the 112i ligand back into the 112i protein structure Load the file into GOLD via GOLD Setup and Runa Docking Load Existing then navigate to the folder containing the tutorial7 files select gold_1 2i_1I2i conf and click Open Run this GOLD job and analyse the results in Hermes to check that the crystallographic binding mode is indeed retrieved Note that once the GOLD run has finished the docking results can be read directly into Hermes from the GOLD Run window via the View Solutions button Read in the file 112i_lig mol2 to make the comparison The GOLD configuration file gold_1x7r_1I2i conf is set up to dock the 112i ligand into the 1x7r protein structure Load the gold_1x7r_1I2i conf as described above run this GOLD job then analyse the results in Hermes Read in the file 1x7r_1 2i_sup mol2 to compare the docked poses with the binding mode found in 112i You may find that there are some solutions which have approximately the right binding mode which return scores of between 23 and 25 However there should also exist higher ranking poses with scores of between 28 and 32 These poses have the ligand rotated thr
262. ngine environment e GOLD can be run in parallel using a Grid Engine environment e Fora Grid computing and Grid Engine overview including documentation and How To guides please see http wiki gridengine info wiki index php Main_Page e For further information and template scripts for running GOLD using a Grid Engine environment please see http www ccdc cam ac uk products life_sciences gold faqs parallel 14 5 2 Using GOLD with Parallel Virtual Machine PVM e GOLD may also be run in parallel via use of PVM Parallel Virtual Machine PVM is a 3rd party public domain library of routines that allows a program to schedule and harvest results across a network of machines and or processors 124 GOLD User Guide e PVM is supplied with GOLD for UNIX based platforms and allows users to distribute jobs over their network across a virtual cluster of machines in order to harness the processing power of multiple machines concurrently e As PVM is now relatively old technology we do not recommend its use and instead recommend use of PBS scripts as detailed in see Using GOLD with a Grid Engine environment However details of how to configure your systems for PVM and how to run GOLD jobs with it are detailed at http www ccdc cam ac uk SupportandResources Support Pages SupportSolution a spx Psupportsolutionid 124 GOLD User Guide 125 15 Viewing and Analysing Results 15 1 15 1 1 15 1 2 126 Description of Output Files Fil
263. nput only the region of interest around the binding site you must ensure that all the residues you include are complete You should also include all residues within a 5A radius from the solvent accessible surface of the cavity Add all hydrogen atoms including those necessary to define the correct ionisation and tautomeric states of residues such as Asp Glu and His see Protonation and Tautomeric States Ensure that all bond types are correct If they are and hydrogen atoms have been placed on the correct atoms GOLD will deduce atom types automatically see Automatically Setting Atom and Bond Types This also applies to PDB input files but only for known residues i e there is no HET group library GOLD connects atoms within residues on the basis of proximity Double bonds are assigned as appropriate for the naturally occurring protein residues Residues should be in sequence order and correctly named All atoms should be properly labelled CA CB etc Any unusual bonds disulphide bridges etc should have CONECT records If a metal ion is present ensure that all bonds between the ion and coordinating protein or water atoms are deleted GOLD will re find them automatically Metals should be within bonding distance of at least two protein and or water atoms in the active site so that GOLD can infer likely coordination geometries see Metal lons Save the protein in e g MOL2 format GOLD assigns atom types from the informati
264. o remove the requirement for fitting points to be solvent accessible see Solvent Accessibility In this case fitting points would be generated for all solvent accessible donor and acceptor atoms within the binding site Remember that these atoms are already deemed to be solvent accessible but it s their potential fitting points that may have been desolvated by neighbouring atoms GOLD User Guide e A Fitting points summary is provided in the gold_protein og file The polar fitting points used by GOLD are also saved as protein atom subsets within Hermes Two subsets are saved donor hydrogens and lone pairs You can highlight the atoms belonging to any subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area 3 6 2 Defining a Binding Site from an Atom e Click on Define Binding Site from the list of Global Options given on the left of the GOLD Setup window e Switch on the button labelled Atom Then within the Hermes visualiser select a single solvent accessible protein atom close to the centre of the active site of the protein 3 GOLD Setup Biel Ez Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Atom select an atom in the visualiser or enter an atom index Proteins Define Binding Site ZN ZN262 View Select Ligands Waters Point
265. o set up a custom square planar geometry you must specify four points using the following vectors 0O 1 0 1 Op 0 1 0 0 0 1 0 GOLD User Guide 23 Assuming the metal is on the origin 0 0 0 GOLD will then attempt to match the specified vectors onto the metal to protein atom vectors found in the protein vectors are normalised to a metal to chelator distance of 2 0 A Once vectors for each point in the polyhedron have been defined click on the Add or replace button to add the custom definition to the list of coordination geometries available for selection see Specifying Metal Coordination Geometries Manually Repeat the above procedure if you want to specify additional custom polyhedron It is possible to set up to three custom metal polyhedron To edit a custom polyhedra highlight the corresponding entry in the The Define custom metal polyhedra window make the required changes and then hit the Add or replace button To remove a custom polyhedra highlight the corresponding entry in the Define custom metal polyhedra window and hit the Delete button or to remove all entries hit the Clear button Once defined the custom geometries will be available for selection when manually specifying allowed coordination geometries see Specifying Metal Coordination Geometries Manually 3 9 5 Metal Ligand Interactions Metal coordination in GOLD is modelled as pseudo hydrogen bonding Metal ligand interactions will typically invo
266. o the trans conformation will also occur if the O C N H torsion is greater than twenty degrees out of plane Note If the O C N H torsion is greater than five but less than 20 degrees out of the trans plane the bond will not be flattened and a warning message will be written to the gold err file Click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Flip amide bonds check box to allow amides thioamides ureas and thioureas in the ligand to flip between cis and trans conformations during docking In order to flip between cis and trans conformations the CO NRR torsion is first made planar at the initialised trans conformation Note N N disubstituted amides are not made planar CO NH will be set so that the NH group is in plane with the CO care must be taken that the input RNH group itself is planar since GOLD will not change this On occasion this flattening of the CO NRR torsion may result in clashes in the initialised structure If this occurs it is advisable to turn off normalisation of amide bonds using the FLATTEN BONDS keyword in the gold params file In this case it is recommended to fix the bond by switching off Flip amide bonds or by explicitly specifying that the appropriate rotatable bonds are held at their input conformation see Fixing Rotatable Bonds at Their Input Conformation If the use of torsion angle distribution has been enabled see
267. obal Options given on the left of the GOLD Setup window and select GoldScore from the Scoring Function drop down menu Then either enter the path and filename of the Scoring function parameter file or click on the button and use the file selection window to choose the file 7 4 ChemScore 7 4 1 Overview 60 The ChemScore scoring function is published in M D Eldridge C W Murray T R Auton G V Paolini and R P Mee J Comput Aided Mol Des 11 425 445 1997 C A Baxter C W Murray D E Clark D R Westhead and M D Eldridge Proteins 33 367 382 1998 ChemScore was derived empirically from a set of 82 protein ligand complexes for which measured binding affinities were available GOLD User Guide ChemScore AG P c 7 4 2 GOLD User Guide binding Unlike GoldScore the ChemScore function was trained by regression against measured affinity data although there is no clear indication that it is superior to GoldScore in predicting affinities ChemScore estimates the total free energy change that occurs on ligand binding as AG AG pona F AGa AG HAG hbond ta Each component of this equation is the product of a term dependent on the magnitude of a particular physical contribution to free energy e g hydrogen bonding and a scale factor determined by regression i e AG 0 0 A G hbond 9 1 Pian A G metal i Me 2 metal AG ipo 3 lipo AG 2 S Per rot 4 rot Here
268. oe Wizard step 3 Define the binding site The binding site can be defined by several different ways an atom a point or a reference ligand Atoms can be selected in the visualiser Global Options 1ACM Wizard steps Atom select an atom in the visualiser or enter an atom index 1 Select a protein 2 Protein setup Vier 3 Define the binding site 4 Configuration template Point select atoms to define a centroid or edit XYZ 5 Select ligands 6 Choose a fitness function aa 7 GA search options 8 Finish One or more ligands c 1ACM List of atoms or residues Select all atoms within 10 A _ Generate a cavity atoms file from the selection Refine Selection Detect cavity restrict atom selection to solvent accessible surface Force all H bond donors acceptors to be treated as solvent accessible Add Definition as a Selection Help R lt Back Next gt Cancel Wizard It is necessary to specify the approximate centre and extent of the protein binding site this can be done in a number of ways from within the Define the binding site window including from a protein atom see Defining a Binding Site from an Atom from a point see Defining a Binding Site from a Point from a reference ligand see Defining a Binding Site from a Reference Ligand from a file containing a list of atoms or residues see Defining a Binding Site from a List of Atoms or Residues For
269. ol2 the log file will be named Myfile rescore log For each rescored ligand a total fitness score and the component scoring terms are listed Status gives an indication of whether or not there were any errors during the rescoring run Simplex indicates whether or not a locally optimised ligand pose was used for the rescoring 1 indicates that the minimised pose was used 0 indicates that the minimised pose was not used and indicates that simplexing was not switched on see Setting Up a Rescoring Run Note When Perform local optimisation simplexing is switched on the minimised conformation will only be used for the rescoring if this results in an improvement to the fitness score When a minimised ligand pose is used for the rescoring an RMSd measure is given of the final minimised orientation with respect to the input ligand conformation The example file below was generated by rescoring the best solution found m2 for the second ligand in the solution file resu ts mol2 Status Simplex RMSd Fitness S hb_ext S vdw_ext S hb_int S int Nr Ligand file Ok 1 0 00 68 72 33 31 25 75 0 00 0 00 1 gold_soln_results_m2_2 molz 15 1 8 Protein Log File The protein log file gold_protein log details the parameterisation of the protein and the determination of the binding site The file is line buffered so you can see how the algorithm is progressing even when GOLD is run in the background GOLD User Guide 129 15
270. om types from the information about element types and bond orders in the input structure file so it is important that these are correct However if for any reason GOLD is unable to deduce an atom type then the atom in question will be replaced with a dummy atom type Du If this is the case a warning message will be given in the gold_ligand log file The presence of dummy atoms should not significantly affect the docking prediction since dummy atoms are neither considered as donors or acceptors There is usually a right and a wrong way to code groups which can be drawn in more than one way i e have more than one canonical form such as nitro carboxylate and amidinium see Atom and Bond Type Conventions for Difficult Groups The starting geometry of the ligand should be reasonably low in energy since GOLD will not alter bond lengths or angles or rotate rigid bonds such as amide linkages double bonds and certain bonds to trigonal nitrogens However GOLD will optimise the values of torsion angles around rotatable bonds Save the ligand as a MOL2 file i e Tripos format or a MOL file i e MDL SD format It is also possible but not recommended to use PDB format If using PDB format CONECT records should also be included see Ligand File Formats Ligand Hydrogen Atoms lonisation States and Tautomeric States GOLD uses an all atom model so the ligand must have all hydrogen atoms added The precise geometrical positions of rotatabl
271. on about element types and bond orders in the input structure file so it is important that these are correct However if for any reason GOLD is unable to deduce an atom type then the atom in question will be replaced with a dummy atom type Du If this is the case a warning message will be given in the gold_protein log file The presence of dummy atoms should not significantly affect the docking prediction since dummy atoms are neither considered as donors or acceptors Note that the steps above are essential whether docking a ligand into a single protein or carrying out an ensemble docking see Ensemble Docking 3 2 Specifying the Protein File or Files GOLD User Guide Click on Proteins from the list of Global Options given on the left of the GOLD Setup window gt GOLD Setup Zox Conf file FN load Save Options 1acm 1a42 107 Wizard Templates Select proteins to use Load Protein Superimpose Proteins Define Binding Site Select Ligands ics Waters 1A42 Ligand Flexibility 1QBT Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Atom Typing Protein score offset ensemble docking only Help R Run GOLD Run GOLD In The Background Finish Cancel e A list of those proteins currently loaded in the Hermes visualiser is available via the Loaded Proteins pull down menu Select the protein you wish to use from this list e Alternatively to specify a di
272. onal argument that instructs the script to use the specified reference ligand file to identify which chain to use in case the unique residue cannot be identified from the chain ID 24 5 3 gold_utils convert e This utility enables file conversion e g pdb to mol2 e Usage gold utils help convert i lt filename gt o lt filename gt e Details of the arguments above are convert required argument that converts the file based on the formats given by the i and o name extensions i required argument that specifies the input molecule file o required argument that specifies the output molecule file 24 5 4 gold_utils write_complexes conf e This file enables a set of protein ligand complex files to be written out for a set of docking solutions e Usage gold utils write_complexes conf lt conf filename gt o lt output_directory gt format lt output_format gt e Details of the arguments above are write complexes required argument that writes all protein ligand complexes for a set of docking solutions o optional argument that specifies the output molecule file format an optional Argument that specifies the output format 232 GOLD User Guide 25 Appendix F The Torsion Angle Distribution File 25 1 Format of Torsion Angle Distribution File Header The first section of the torsion angle distribution file sets parameters and tells GOLD what to do with the distributions
273. onal information will also be provided on the handling and parameterisation of metals in GOLD Preparation of Input Files GOLD User Guide Open Hermes and read in the file protein mol2 from the folder to which you copied the tutorial2 files The original PDB file 1442 pdb has also been provided should you wish to set up the protein for yourself The protein protein mol2 has already been set up in accordance with the guidelines for the preparation of protein input files see Setting Up the Protein s Upon inspection of protein mol2 you should notice that parts of the protein remote from the binding site have been deleted in order to speed up the calculation see Using the GOLD Wizard to Prepare the Protein File You should also notice that all water molecules have been deleted and that hydrogen atoms have been placed on the protein in order to ensure that the ionisation and tautomeric states are defined unambiguously see Protonation and Tautomeric States There are some additional requirements when preparing a protein input file which contains a metal ion Inthe original protein file it is essential that the metal ion is coordinated to at least two protein atoms or water molecules so that GOLD can determine the correct coordination geometry Inthe prepared protein input file the metal ion must not have any bonds to coordinating atoms If these are present in the original protein file they must be deleted On closer inspect
274. or more motifs can be specified and each motif will consists of a unique combination of interactions formed between the protein and the ligand Individual interactions are described according to their protein atom interaction type hydrogen bond acceptor hydrogen bond donor lipophilic interaction or weak CH O acceptor GOLD assesses whether or not specified interactions are satisfied as follows For Hbond acceptor and Hbond donor interaction types A hydrogen bond is deemed to be present if the distance between the acceptor and the donor heavy atoms are within the range 2 85A 0 45 the acceptor angle is within 1450 65 and the donor angle is within 115 40 Further a planarity check is used to ensure that the hydrogen bond is not more than 30 out of the plane when the ligand donor is of atom type N 2 or N pl3 or the protein donor is of atom type N pl3 or N am Donor Angle NAY C Acceptor Angle 103 10 7 2 104 For a weak CH O acceptor interaction type an interaction is deemed to be present if the distance between the acceptor and the aromatic carbon is within the range 3 35A 0 65 the acceptor angle is within 145A 65 the donor angle is within 115A 40 and the CH O bond is not more than 30 out of the aromatic plane Note the presence of a heteroatom X below where X is O 0 3 N N ar N 2 N am N pl3 S S 3 is required in the aromatic ring Ox acceptor Ge ea angle SF For a lipo
275. ormalise the position of H atoms bonded to other elements For further information refer to the Hermes user guide The geometry of added hydrogen atoms will be chemically meaningful However the precise geometrical positions of Ser Thr and Tyr hydroxyl hydrogen atoms or Lys NH3 hydrogen atoms do not matter as their orientation will be optimised during the GOLD run see Rotatable O H and NH Groups GOLD deduces the hydrogen bonding abilities of protein residues from the presence or absence of hydrogen atoms For example you can control the protonation and tautomeric state of Asp Glu and His residues by adding or removing appropriate hydrogen atoms If incorrect ionisation or tautomeric states are inferred by the program it is unlikely that correct protein ligand binding modes will be predicted It is therefore important that you check protonation states of such residues before proceeding with the docking Additional structure editing functionality is available within the Hermes visualiser 3 3 2 Applying Protonation Rules It is possible to protonate using SMARTS based protonation rules contained in a protonation_rules txt file A sample file is provided in lt install_dir gt GOLD Suite Hermes The file contains SMARTS based rules for protonation of the format lt query SMARTS gt lt rule SMARTS gt e g carboxylat set the C O bond type to aromatic C OD OD C OD OD Load this file using the file selection
276. ough 180 degrees along the long axis as shown in the superposition below crystallographic binding mode colour coded orange GOLD docking pose colour coded green 195 Cross Docking into 1x7r with a Soft Potential applied to Leu 346 196 Load the file gold_1x7r_1I2i_SP conf into Hermes The definition of soft potentials is specific to the protein thus click on the MOL_ID 1 tab adjacent to the Global Options tab Click on Soft Potentials You will notice that LEU346 is already in the Residues alternative potential 1 box This means that a soft vdW potential with 2 4 functional form has been applied to one residue only Leu346 This replaces the default 4 8 functional form that applies to the rest of the protein Note Potentials are applied simply by choosing the appropriate potential i e either 1 or 2 and activating the Add selections radio button adjacent Residues are then added to the appropriate box by selecting them in the 3D view Because LEU346 is in the Residues alternative potential 1 box this means a 2 4 soft potential is applied to the residue If LEU346 had been entered into the Residues alternative potential 2 this would mean a softer 1 2 functional form would be applied Further information is available see Allowing For Localised Movements Docking With Soft Potentials Run the docking job gold_1x7r_1I21_ SP conf and analyse the results using Hermes This time you should find that the highest scoring solut
277. params HBOND_F Hydrogen bonding parameters for potential H Bond A to D are distance parameters E and F are interaction scores BURIED_A BURIED_B BURIED_C BURIED_D BURIED_E BURIED_F Parameters used for potential buried A to D are distance parameters E and F are interaction scores METAL_A METAL_B METAL_C METAL_D METAL_E METAL_F Metal bonding parameters for potential metal A to D are distance parameters E and F are interaction scores NONPOLAR_A NONPOLAR_B NONPOLAR_C NONPOLAR_D NONPOLAR_E NONPOLAR_F Parameters for potential nonpolar A to D are distance parameters E and F are interaction scores REPULSIVE_A REPULSIVE_B REPULSIVE_C REPULSIVE_D Parameters for potential repulsive A and B are distance parameters C and D are interaction scores LINK_BEND_COEFFICIENT see ChemScore GOLD User Guide lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt lt float value gt 57 58 Parameters for additional ChemScore contributions used in CHEMPLP chemplp params
278. pecified See plp params and chemplp params for the default parameters used Figure Piecewise Linear Potential left Partially attractive potential right Repulsive potential Protein atom type Ligand atom type donor Acceptor don acc nonpolar metal donor repulsive H bond H bond buried repulsive acceptor H bond Repulsive H bond buried metal don acc H bond H bond H bond buried metal nonpolar buried Buried buried nonpolar buried Table PLP interaction types selected depend on the protein and ligand atom type GOLD User Guide 55 7 2 3 56 Altering PLP Fitness Function parameters The PLP and CHEMPLP parameter files are stored in the GOLD_HOME gold directory They contain all the parameters used by the GOLD implementation of PLP A full description of the meaning of the PLP specific parameters are given below The PLP and CHEMPLP files can be customised by copying them editing the copy and instructing GOLD to use the edited file To use a modified plp params or chemplp paranms file click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select PLP from the Scoring Function drop down menu Then either enter the path and filename of the Scoring function parameter file or click on the button and use the file selection window to choose the file The format of the PLP and CHEMPLP file is quite strict incorrect editing may cause GOLD to behave in unexpected w
279. penalising poses that do not fulfil a specific constraint As such if a constraint is only set up on a subset of all models the proteins without constraints could end up being favoured over those with the constraint In addition it is worth noting that docking the same ligand into different protein models can lead to differences in the scores Thus in order for the constraints to have a noticeable effect one might need to increase their weighting from the default values The combination of constraints and ensemble docking is not a straightforward problem care should be taken in order to obtain results that are meaningful 4 4 Allowing For Localised Movements Docking With Soft Potentials GOLD User Guide GOLD uses Lennard Jones functional forms for both the External and Internal Van der Waals contributions to the Fitness Function By default a 6 12 potential is applied to the Internal Van der Waals contribution and a 4 8 potential is applied to the External Van der Waals contribution These defaults are defined in the gold params file see Altering GOLD Parameters the gold params File The 4 8 potential form for the External contribution is selected as being optimum for general use However there are cases where this potential form may be too severe in the short contact i e the clash component This would arise for instance where part of the binding site is made up of a loop which it is known can move aside 37 38 slightly to accommod
280. philic interaction type an interaction is deemed to be present if the sum of the protein and the ligand atom s van der Waals radii plus 0 4 is less than the distance between the protein and the ligand atoms During docking a contribution will be added to the fitness score of ligand poses in which a motif is matched i e poses in which all the interactions defined as part of a motif are satisfied This contribution is based upon the accumulated hydrogen bonding and lipophilc interactions defined as part of that motif Therefore docking will be biased towards ligand poses which form interactions to the protein atoms of interest matching one of the uniquely defined motifs Please note that it is not possible to manipulate interaction motif constraints within the constraint editor see Using the Constraint Editor Setting up an Interaction Motif Constraint To define an interaction motif constraint click on Interaction Motif from the list of Global Options given on the left of the GOLD Setup window One or more motifs can be specified and each motif will consists of a unique combination of interactions formed between the protein and the ligand To define an interaction Click on the Add Interaction button Interactions are described according to their protein atom interaction type Select the type of interaction hydrogen bond acceptor hydrogen bond donor lipophilic interaction or weak CH O acceptor using the drop down menu under
281. protein flexibility In this example GOLD will be used to perform a non native docking of a cyclic nucleotide phosphodiesterase 5A PDESA inhibitor the inhibitor from the PDB entry code 1x0z into an ensemble of four PDESA protein conformers PDB entry codes 1xp0 1t9s 1tbf and 2chm in order to investigate how this inhibitor molecule fits into non native proteins from the same family The PDESA proteins cleave phosphodiester bonds in the second messenger molecules cyclic adenosine monophosphate and cyclic guanosine monophosphate PDESA inhibitors are mainly used as treatments for erectile dysfunction and pulmonary hypertension The input files have been taken from the Astex Non native Set More information can be found in the following case study on our website http www ccdc cam ac uk lists resourcefilelist ensemble_docking pdf This tutorial is intended to give an overview to illustrate how ensemble docking is set up The set up is similar to that of a standard docking but there are some aspects that are specific to ensemble docking This tutorial assumes some prior knowledge of GOLD and setting up of dockings Please refer to the standard GOLD tutorials in the GOLD documentation if you do not have this prior knowledge of docking with GOLD Getting Started GOLD User Guide First copy the files in lt install_dir gt GOLD Suite GOLD examples tutoriall0 toa directory to which you have write permissions Open Hermes Open the GOL
282. protein you will see that the zinc atom is coordinated to three histidine groups the one remaining zinc coordination site is available for binding to the ligand e Read in the file ligand_reference mol2 from the folder to which you copied the tutorial 4 files Inspect the crystallographically observed position of the ETS inhibitor shown in green within the protein binding site 172 GOLD User Guide The terminal sulphonamide nitrogen atom of the ligand clearly coordinates to the zinc We can attempt to reproduce this known binding mode within GOLD with the introduction of a distance constraint during docking Ten ligands each structurally similar to the ETS inhibitor will be screened using GOLD These ligands were identified using Relibase a program for search and analysis of protein ligand complexes http www ccdc cam ac uk products life_sciences relibase These ligands ligand mol2 are available from the folder to which you copied the tutorial 4 files note that each of the ten ligands in this file features a terminal sulphonamide group If you have opened all of the files above close them by going to File Close All Files A configuration file gold conf has been provided for this tutorial which will automatically load the settings and parameter values for this tutorial into the GOLD front end From within Hermes click on GOLD then Setup and Run a Docking in the top level menu Load the gold conf for tutorial 4 by selec
283. pt GA button in the Run GOLD window to interrupt and terminate the docking run e Once the job is complete the docked ligand solutions can be viewed in the Hermes visualiser To do this click on the View Solutions button in the Run GOLD window GOLD User Guide 123 14 3 Submitting a GOLD job to the Background e You can submit a GOLD job the background by using the Run GOLD in background button in the GOLD Setup window having first specified all the required information such as protein and ligand file names parameter settings etc 14 4 Running GOLD from the Command Line e Unix platforms GOLD can be run directly in the background by using a simple command available in GOLD_DIR bin gold auto gold conf amp where gold conf is the name of a configuration file e Windows Platforms GOLD can be run on Windows by starting a command prompt navigating to the directory containing the gold conf file and running the following command C Program Files CCDC GOLD_Suite GOLD gold d_win32 bin gold_win32 ex en The above command assumes that GOLD is installed in the default installation directory and that the configuration file is called gold conf If another name has been used for the gold conf e g new_conf_filename conf this will have to be specified C Program Files CCDC GOLD_ Suite GOLD gold d_win32 bin gold_win32 ex e new conf filename conf 14 5 Running in Parallel 14 5 1 Using GOLD with a Grid E
284. put structure file so it is important that these are correct see Atom and Bond Type Conventions for Difficult Groups However if for any reason GOLD is unable to deduce an atom type then the atom in question will be replaced with a dummy atom type Du It does not matter whether the bonds in an aromatic ring are coded as aromatic ar or alternate single and double as the GOLD atom type assigner will automatically assign the special SYBYL bond type ar where appropriate The atom type assigner will also detect amide linkages and assign them the SYBYL bond type am 49 6 3 50 Care should be taken when using the type assignment software on protein input files In particular the software is likely to be unreliable if protein residues have been partially deleted so that some atoms appear to have free valencies This situation can be avoided by ensuring that all residues included in the input file are complete There is usually a right and a wrong way to code groups which can be drawn in more than one way i e have more than one canonical form such as nitro carboxylate and amidinium A list of correct bond types for some of the common difficult groups is available see Atom and Bond Type Conventions for Difficult Groups Because correct atom typing is so important any messages from the type checker are logged in both the gold_protein log file and the gold err file These errors will also be displayed in a separate window if GOLD
285. raint weight is also the value of the penalty applied to the fitness score for each constrained H bond that is not formed The Minimum H bond geometry weight is a user defined score that determines how good a hydrogen bonding interaction has to be in order for it to be considered a hydrogen bond by GOLD The Minimum H bond geometry weight takes a range of values from 0 to 1 by default this value is set at 0 005 For a given protein H bond constraint more than one protein atom number can be entered in the Protein atom s required to form H bond entry box This will instruct GOLD to use an either or type of constraint during docking For example specifying two protein atoms acceptor m and acceptor n will result in the constraint being satisfied if an H bond is formed to either m or n during docking This is of use when defining constraints involving for example carboxylates where it is not important which oxygen atom forms an H bond provided that one of them does Click on the Add button to add the constraint definition to the constraint editor see Using the Constraint Editor It is possible to specify several different protein H bond constraints with different weights for each constraint 97 10 4 Region Hydrophobic Constraints e This constraint can be used to bias the docking towards solutions in which particular regions of the binding site are occupied by specific ligand atoms or types of ligand atom e g hydrophobic atoms 1
286. ranked pose when compared to that of the co crystallised ligand is 7 30 A Clear up the 3D view by selecting File then Close All Files Running a Diverse Solutions Docking and Viewing the Results 200 The docking set up is provided in diverse conf and the corresponding output is in the diverse directory in the folder to which you copied the tutorial8 files The docking results can be read into Hermes via File Load GOLD results Alternatively the diverse solutions docking can be set up by modifying the conf file from the previous exercise instructions on how to do this follow The GOLD interface will still be open Click on the Load button to read the gold conf from above Click on Fitness amp Search Options then activate the Generate Diverse Solutions tickbox The settings for generating diverse solutions can be modified click on the Diverse Solutions Options button to view these settings Change the Cluster size to 2 and the R M S D to 2 0 Angstroms This means that each diverse solutions cluster will contain 2 ligands and that the clusters will differ by an RMSD of 2 A see Setting Up GOLD to Generate Diverse Solutions Hit Close to close the window Now click on Output Options and change the directory from what it was previously to e g diverse solutions At the bottom of the window activate the Create links for different binding modes based on RMSD clustering tick box then enter 2 0 into the box next to Distance between clus
287. re covered in the sections that follow If you wish you can run the two GOLD jobs using the configuration files provided Alternatively you can view the results that we have generated Since GOLD is non deterministic any results that you get might differ from ours but the general trends are likely to be the same Running the Non flexible Docking and Analysing the Results GOLD User Guide Open Hermes and specify the non_flexible conf via GOLD Setup and run a Docking Load Existing Click on the Protein 1fax coagulation factor tab then click on the Flexible Sidechains option The defined active site in the protein has been broken down into its constituent residues which are provided in a scrolling list Scan through the list you will notice that the Status of all the residues is listed as Rigid Return to the Global Options tab where we can define general docking settings and click on Output Options Change the output directory name e g to non_flexible2 then click on Run GOLD When prompted that The GOLD configuration has been updated and needs saving click OK then change the GOLD conf file name e g to non_flexible2 conf Start the docking by clicking on Save Once the docking has completed load the solutions into Hermes using the View Solutions button then Close to close the Run GOLD window Load the 1fax_ilpg_super mol2 superposition file into Hermes via File Open Hit the Display tab in the Molecule Explorer section of the
288. rical or text information that is present as tagged fields in the sdf or mol2 files used to create the GoldMine DB Such data may include the individual terms that make up the scoring function used in the docking see Controlling the Information Written to Ligand Solution Files Each individual quantity for which a set of data is saved is termed a Descriptor Additional descriptors calculated using Hermes can also be added to a GoldMine DB and used in further analysis see Viewing Docked Solutions in Hermes Numerical and text data for descriptors associated with individual poses can be viewed and sorted within a spreadsheet Histograms and scatter plots can be generated for any numeric descriptors GoldMine allows you to filter your results in a sophisticated manner Ranges for a number of descriptors can be set and the ranges combined in Boolean fashion to create sets of docking poses satisfying appropriate properties Statistical analysis and modelling can be carried out and new scoring functions derived Test Virtual Screening runs can be analysed using ROC curves For further information refer to the GoldMine user guide 15 6 2 Sending Docking Results to GoldMine GOLD User Guide Docking results can be sent to GoldMine and saved within a GoldMine database The docked poses are saved within GoldMine in a new or existing dock set and the protein file used in the docking is the protein model associated with that dock set Fir
289. rnk files check box 15 1 5 File Containing Ranked Fitness Scores for a Set of Ligands e Afile called bestranking 1st is written for batch jobs on multiple ligands This gives a continuous summary of the best solution that has been obtained for each completed ligand e To specify an alternative filename click on Output Options from the list of Global Options given on the left of the GOLD Setup window then select the File Format Options tab Enable to Use alternative bestranking I st filename check box and either enter the new path and filename or click on the button and use the file selection window to choose the file e The file gives total fitness scores and a breakdown of the fitness into its constituent energy terms The example file below was generated from a ligand input file containing 5 ligands The listed file names correspond to the names of the files containing the best solution found for each ligand e g gold soln ligs m1 3 mol12 contains the best answer found for the first ligand in the input file File containing a listing of the fitness of the top ranked individual for each ligand docked in GOLD Format is Fitness S hb_ext S vdw_ext S hb_int S int File name Ligand name 51 93 13 62 32 46 0 00 6 31 home GOLD gold_soln_ligand_m1_3 mo12 LIN AL5_555_pdbibni_1 43 80 11 47 28 91 0 00 7 42 home GOLD gold_soln_ligand_m2_8 mo12 LIM 4L1_555_pdbibnn_1 42 56 7 38 31 74 0 00 8 46 home
290. rogen bonding to the ligand Lysine NH3 groups are similarly optimised GOLD can allow side chains to rotate within user defined bounds during docking see Side Chain Flexibility GOLD can dock into multiple conformations of the same protein see Ensemble Docking Note that the final positions of any movable protein atoms that are generated during docking these will usually be different for each docked ligand pose can be saved to the docked solution file see Controlling the Information Written to Ligand Solution Files e Ligand flexibility Only the torsions around the ligand s flexible bonds will be optimised during docking Bond distances and valence angles must be optimised before using GOLD Torsion angle distributions extracted from the Cambridge Structural Database CSD can be used to restrict the ligand conformational space sampled by the genetic algorithm Using torsion angle distributions in this way may improve the chances of GOLD finding the correct answer by biasing the search towards ligand torsion angle values that are commonly observed in crystal structures It may also improve convergence and so make GOLD usable with faster settings see Enabling Use of Torsion Angle Distributions The use of torsion angle distributions is enabled by default The torsion distribution file go d tordist is provided in lt install_dir gt GOLD Suite GOLD gold and can be manually edited to include specific user defi
291. rs can be exploited using receptor depth scaling where 116 GOLD User Guide the score attributed to hydrogen bonds is scaled depending on the depth in the pocket Hydrogen bonds deep in the pocket are rewarded with an increased score while the scores of those closer to the solvent exposed surface are decreased Simultaneously the scores attributed to lipophilic interactions are reduced This procedure has been shown to increase the relative scores of active molecules compared to inactive molecules across a diverse range of 85 proteins see References O Boyle Brewerton and Taylor While the default values for the parameters are suitable for the general case for a particular protein it may be possible to gain better results by adjusting the scaling parameters see the GOLD configuration file documentation for further information Simplexing is turned on when rescoring with receptor depth scaling Receptor depth scaling is only available when rescoring with ChemScore see Setting Up a Rescoring Run 12 5 Rescore Output Files GOLD User Guide Each rescored solution is written to the rescore 1og file This file contains the ligand identifiers the final rescore fitness value and its component terms To specify an alternative rescore log filename see Rescore Log File Rescored structure solution files can be written out that will contain the new scoring function terms and can be used with GoldMine Solutions will be written to th
292. ructure is number 1 in the molecule input file and the solution is from the fourth docking dock4 The format for the output of the equivalent sd input file would be the following ligand sd 1 dock4 To revert to the historic output i e to output only the structure name e g N phosphonacetyl L aspartate the line SET UNIQUE SOLN TITLES 1 inthe gold params file should be changed to read SET UNIQUE SOLN TITLES 0 File Containing the Protein Binding Site Geometry During docking GOLD will optimise hydrogen bond geometries by rotating groups such as serine OH and lysine NH3 It is also possible to allow specific protein side chains to be treated as flexible during docking see Side Chain Flexibility This means that the coordinates of certain protein atoms such as these will change Protein atom positions that are generated during docking will usually be different for each docked ligand pose and are therefore written to the individual ligand solution files This information can be written to SD file tags for MOL2 files these tags are written to comment blocks see Controlling the Information Written to Ligand Solution Files Structure files containing the optimised protein binding site geometry can be written out from the Hermes visualiser File Containing Ranked Fitness Scores for an Individual Ligand A file called lt ligand_file name gt _m rnk is written for each ligand m refers to the position o
293. ry 1 F Custom geometry 2 F Custom geometry 3 All None Define Custom Polyhedra Set Default Delete No custom polyhedra Click on the metal atoms in the above list to highlight the coordinating atoms and to set the coordination geometry Help R Run GOLD Run GOLD In The Background Finish Cancel Any metals in the currently loaded protein will be recognised and listed By default only the coordination geometries for the corresponding metal type defined in the gold params file will be considered during docking For example for a Zn atom GOLD will attempt to match coordination geometries 4 5 and 6 tetrahedral trigonal bipyramidal and octahedral templates onto the coordinating atoms found in the protein see Automatic Determination of Metal Coordination Geometries If you wish to manually specify coordination geometries for particular metal atoms then select the allowed coordination geometries by enabling the corresponding check box es Once the allowed geometries have been selected for a particular metal atom click on the Set button If the list of pre defined coordination geometries does not contain a suitable geometry then you can define a custom metal coordination geometry see Defining Custom Metal Coordination Geometries GOLD User Guide 3 9 4 To return the allowed coordination geometries of a particular metal to the defaults defined in the gold params file highlight the entry and hit the De
294. s GOLD proceeds symbolic links are created ranked_structure_m _1 mol2 will always point to the current top ranked solution ranked_structure_m 2 mol2 will point to the second best solution and so on Alternatively you can specify that all saved docking solutions for all ligands are to be concatenated and written to a single file see Specifying Ligand Solution File Formats and Directories Output files for the docked ligand s may also contain additional information such as the scoring function terms and the rotated protein hydrogen atom positions specific to that solution This information can be written to SD file tags for MOL2 files these tags are written to comment blocks It is possible to control the information written to solution files see Controlling the Information Written to Ligand Solution Files A description of the various other tags available can be found in see Appendix C Additional Tags in Output Files Solution file title strings take the form lt file basename gt lt p gt cov lt r gt dock lt q gt where lt file _basename gt is the base name of the ligand input file GOLD User Guide 15 1 3 15 1 4 GOLD User Guide lt p gt is the molecule number in the file lt q gt is the number of the docking lt r gt is the covalent attachment atom This part is only printed for covalent dockings For example mol file ligand mol2 1 dock4 where the ligand filename is 1igand mol12 the st
295. s i e a multi MOL2 or SD file it is possible to only dock specific ligands in that file Specify which ligand you wish to start and finish docking at by entering in the First Ligand and Last Ligand boxes the numbers relating to the position of the ligands within the file Unless specified otherwise GOLD will by default start at the first ligand and finish at the ast ligand in the file e Repeat the above procedure if you want to select further ligands for docking e To edit a specified ligand file e g to change the number of times the ligand will be docked highlight the ligand file with the mouse and make the required change e To remove a specified file from those listed highlight the ligand file with the mouse and hit the Delete button e Itis also possible to retrieve and dock ligands directly from a GoldMine database see Receiving Ligands From GoldMine e Itis possible to supply to GOLD with a file containing a reference ligand e g a crystallographically observed ligand pose The ligand reference file will be used to perform automated RMSd calculations against GOLD solution s see Specifying a Ligand Reference File GOLD User Guide 5 6 Receiving Ligands From GoldMine e It is possible to take a selection of ligand poses from a GoldMine analysis and submit them to GOLD for docking The results can then be saved within output files or alternatively they can be returned to GoldMine and saved within a GoldMine DB see Sen
296. sary in the above experiments As long as it can adopt the native 1fax position and one other position in which it is folded away from the binding site that might well have been enough GOLD User Guide One problem is that in some conformations GIn192 tends to clash with Arg143 At first sight this means we have to be careful to pick a GIn192 rotamer that is folded away from the binding region but also does not clash with the arginine residue A way round this is to add the command penalise_protein_clashes 0 to the rotamer_lib command block place it anywhere between rotamer_lib and end_rotamer_lib This will switch off calculation of clashes between flexible side chain atoms and neighbouring protein atoms allowing GlIn192 to approach nearby residues closely While physically unrealistic this is a pragmatic tactic that might well work and is not as egregious as it sounds since in reality Arg143 can probably move away from Gln192 if it needs to Obviously you can experiment with these options if you wish This ends the tutorial GOLD User Guide 193 20 7 Tutorial 7 Docking using Localised Soft Potentials Introduction First copy the files in lt install_dir gt GOLD Suite GOLD examples tutorial7 toa directory to which you have write permissions The object of this tutorial is to demonstrate how to employ the Localised Soft Potential option that is available when using GoldScore This option allows you to soften the
297. score dllor chemscore d11 Windows On UNIX the file 1ibgold so is included in the GOLD distribution together with two versions of libfitfunc_dll so one implementing the normal GOLD scoring function and the other implementing the ChemScore function On Windows the file gold d11_ is included in the GOLD distribution together with two files called goldscore d11 for implementing the normal GOLD scoring function and chemscore d11 for implementing the ChemScore function It effectively provides a mechanism by which data may be intercepted and modified during docking Users may therefore post process the results of a docking or modify the GOLD function or implement their own scoring function by building their own versions of libfitfunc_dll so UNIX or e g goldscore d11 Windows Altering GOLD Parameters the gold params File The parameter file gold params is stored in the GOLD distribution directory It contains all of the parameters used by GOLD e g hydrogen bond energies atom radii and polarisabilities torsion potentials hydrogen bond directionalities etc other than those which are specified in the configuration file i e can be set via the GOLD front end It also contains parameters that control the general behaviour of GOLD e g whether the final solution from a genetic algorithm run is to be minimised via a Simplex procedure before being saved The parameter file can be customised by copying it editing the copy a
298. scoring functions and considering the best results from each can have a favourable impact on the overall rank ordering of ligands see Rescoring User Defined Score allows users to implement their own scoring function or modify an existing scoring function by specifying a path to a dynamically loadable shared object library see User Defined Scoring Function To select a scoring function click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window and select the required scoring function from the drop down menu Piecewise Linear Potential CHEMPLP Overview For a more detailed description of the PLP and CHEMPLP fitness functions as well as the derivation of their parameters please see O Korb T StYtzle and T E Exner see References PLP and CHEMPLP are empirical fitness functions optimised for pose prediction CHEMPLP is the default scoring function in GOLD In both cases the Piecewise Linear Potential fp p is used to model the steric complementarity between protein and ligand while for CHEMPLP additionally the distance and angle dependent hydrogen and metal bonding terms from ChemScore are considered fehem hb Schem chor fchem met The internal score of the ligand consists of the heavy atom clash potential ffig ciasn see References as well as the torsional potential used within ChemScore fiig tors Both fitness functions are capable of covalent docking fcrem
299. se to an extra Protein Energy term which contributes to the total fitness value The term is computed by summing the van der Waals interactions of all pairs of protein atoms which satisfy the following conditions a at least one of the protein atoms is in a flexible side chain b the van der Waals term for that pair of atoms is repulsive The van der Waals interactions will be estimated using the same potential as is used for the protein ligand vdw term by default this is a 4 8 potential The protein protein clash term can be switched off by including the command penalise_protein_clashes 0 anywhere in a rotamer_lib block within the gold conf file For further instructions refer to the GOLD configuration file documentation Note that this will switch off calculation of the protein protein clash term for all flexible side chains not just the one corresponding to the rotamer_lib block in which you have placed the penalise_protein_clashes 0 command GOLD User Guide It is recommended that the protein protein clash term be switched off in the following cases When fixed conformers are used i e delta 0 throughout Fixed conformations will usually arise from having identified all such conformers in different crystal structures and so will be comparable in energetics You may therefore wish to treat each on equal merit They may exhibit different protein protein clash terms however and if this option is switched on one may be signifi
300. se and Astex Statistical Potential scoring functions and the Diverse Solutions code within GOLD 2001 2015 Astex Therapeutics Ltd All rights reserved Licences may be obtained from CCDC Software Ltd 12 Union Road Cambridge CB2 1EZ United Kingdom Web www ccdc cam ac uk Telephone 44 1223 336408 Email admin ccdc cam ac uk GOLD User Guide Contents 1 Vandaele Le e d1 AVAA n E e E er riteesteaviee tessa aeereet aeoat ade at adits 1 2 Getting Started 00 cccccccccccceeceaeeseeeeeeeeeesseeeeeeeeeeeeeseeeeseeeeeeeeessseeeseeeeeeeessaaaaaeeeeeeeeeaaa 2 2 1 Overview of the GOLD Interface eee ee ecscecceeeeeeeeenaeeeeeeeeeeeeeeaaaeeeeeeeeeeeeeaaaaes 2 2 2 Using the GOLD Docking WiZaId ccccccssssssseeccceccaeaeseseeceeeeeeaaaeeeeeeeeeessaaagess 3 3 Setting Up the Proteins is cccccceciceckceckcscheceheeetenetehecenedededeaedegedegedededeuadededeaedeaadeaedeaedenedens 5 3 1 Essential Step iarann canted Hoa awd od aA AeA AAA AeA AAA AAA 5 3 2 Specifying the Protein File Or FileS cccccccccccsessseseecceeeesaeeeseeeeeeeeesssaaaeeeeeeess 5 3 3 Protonation and Tautomeric States eeeeccsceeeeeeeeeeeecaeeeeeeeeeeeeenaaaeeeeeeeeeeees 6 3 3 1 Adding Hydrogen Atoms to the Protein Using Program Defaults 6 3 3 2 Applying Protonation Rules cccssscscccececcesesseeeeeeeeesseeaeseeeeeeeeeeaaa 7 3 3 3 Flipping Asn and Gln R SIGUGS ccccccccsesseseeeeeeeecsaaeeeeeceeeeseaaaeness
301. sed towards fitter members of the population i e chromosomes corresponding to ligand dockings with good fitness scores e Anumber of parameters control the precise operation of the genetic algorithm viz Population size see Population Size Selection pressure see Selection Pressure Number of operations see Number of Operations Number of islands see Number of Islands Niche size see Niche Size Operator weights migrate mutate crossover see Operator Weights Migrate Mutate Crossover Van Der Walls and hydrogen bonding annealing parameters see Van der Waals and Hydrogen Bonding Annealing Parameters e Changes to individual genetic algorithm parameters should be made with care see Using User Defined Genetic Algorithm Parameter Settings 11 3 2 Relationship between Genetic Algorithm Parameters and Speed e The time taken by GOLD to dock ligands can be controlled by altering the values of the genetic algorithm GA parameters e GOLD runs for a fixed number of genetic operations crossover migration mutation The easiest way to make GOLD go faster is to reduce the number of GA operations performed in the course of a run This is done through the Number of Operations variable this parameter is called maxops in the configuration file e Areduction in Number of Operations is likely to change the optimum values of several other GA parameters particularly popsize van der Waals and Hydrogen Bonding e GOLD
302. site Scaffold Region Atom Typing T Never dock a ligand when a constraint is physically impossible Fine ota Help R Run GOLD Run GOLD In The Background Finish Cancel e For all constraints the constraint editor is present at the bottom of each constraint setup page Once the settings for a constraint have been specified click on the Add button to add the constraint definition to the list of defined constraints Repeat this procedure if you want to specify additional constraints e To edit a constraint highlight the corresponding entry in the list make the required change and then hit the Add button e Toremove a constraint from the list highlight the entry and hit the Delete button or to remove all defined constraints hit the Clear button e Itis possible to instruct GOLD not to dock ligands when the specified constraint is physically impossible to satisfy e g if no suitable group is present in the ligand to form the required H bond constraint This is done by selecting the Never dock a ligand when a constraints is physically impossible check box e When using constraints GOLD will be biased towards finding solutions in which the specified constraint is satisfied However it is important to remember that such a solution is not guaranteed i e it is not possible to force a constraint to be satisfied in the final solution 10 2 Distance Constraints e Distance constraints are applicable to individual pro
303. speaking the numbers correspond to the order in which the proteins are loaded This index number is given in the docking solutions pane in Hermes as the column headed Ensemble ID next to the fitness score so you can see to which protein each solution corresponds GOLD gives best docking results for proteins 1 and 4 1t9s and 2chm respectively Protein 2chm gives the best docking results which is not entirely surprising given that it is the most structurally similar protein to 1xoz protein More analysis of the results is given in the corresponding Use Case that can be found on our website http www ccdc cam ac uk lists resourcefilelist ensemble_docking pdf The crystallographically observed conformation of the docked 1xoz_ligand can be compared to the poses found when docking this ligand to the ensemble of PDESA proteins The best ranking pose obtained in protein model 4 i e 2chm is shown below coloured by elements with green carbon atoms It is very close to the crystallographically observed pose for the same ligand in 1xoz shown coloured by elements with pink carbon atoms This ends the tutorial GOLD User Guide 219 21 Appendix B List of Atom and Bond Types GOLD uses SYBYL atom and bond types as follows Atom types Hydrogen H Carbon sp3 C 3 Carbon sp2 C 2 Carbon sp C 1 Carbon aromatic C ar Carbocation guanadinium C cat Nitrogen sp3 N 3 Nitrogen sp2 N 2 Nitrogen sp N 1 Nitrogen aromatic e g in pyridine N ar Ni
304. ss score and also the constituent scoring terms will be written Save per atom scores to charge field Enable this check box to write to the mol2 file charge field of docked solution files the scoring contribution of individual ligand atoms Selecting Which Ligand Solutions to Keep By default GOLD will dock each ligand 10 times starting each time from a different random population of ligand orientations see Number of Dockings This can produce a lot of output and you may therefore wish to reduce the number of docking solutions that are retained Click on Output Options from the list of Global Options given on the left of the GOLD Setup window then select the Select Solutions tab GOLD User Guide 3 GOLD Setup BE Ea Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial9 gald conf Load Save Options Protein 1ase aminotransferase Wizard Templates Proteins Define Binding Site Ligands Keep the best E solutions For each ligand aters Ligand Flexibility Keep the top ranked solutions for the best l 100 ligands only Fitness amp Search Options GA Settings Output Options J Reject solutions with a fitness score lower than foo GoldMine Parallel GOLD Constraints Atom Typing File Format Options Information in File Selecting Solutions Keep all solutions E Help h Run GOLD Run GOLD In The Background Finish Cancel e By selecting the appropriate option it is
305. ssible to define multiple constraints e g one for donors and one for acceptors 10 6 Scaffold Match Constraint e The scaffold match constraint can be used to place a fragment at an exact specified position in the binding site the geometry of the fragment will not be altered during docking 10 6 1 Method Used for Scaffold Match Constraint e This constraint will attempt to a place a ligand onto a given scaffold location The scaffold can for example be a common core or fragment useful when docking ligands of a combinatorial set or it may just be a substructure known to adopt a certain binding position e Itis advised that only those atoms required for scaffold matching are specified when using the scaffold constraint Having a scaffold that almost exactly matches the docked ligand and specifying a large number of atoms for matching causes GOLD problems when it is generating random and unique individuals during docking GOLD User Guide 101 10 6 2 102 The scaffold must be supplied as a mo12 file The file should contain the scaffold fragment in its docked position i e expressed in the same coordinate frame as the protein and with the coordinates required to place it in the correct pose The element type is matched not the atom type thus it is not essential for the SYBYL atom types to be correct in the scaffold mol2 file It is recommended that the scaffold have hydrogens correctly placed on all appropriate atoms other tha
306. st it is necessary to create or load a valid GOLD configuration file Since it will often be the case that you will be using a tried and tested docking protocol to redock selected ligands reading in an existing file is probably the usual case Click on GOLDMine from the list of Global Options given on the left of the GOLD Setup window Within this pane it is possible to set GOLD up so that it receives and or sends docking poses from GoldMine 135 3 GOLD Setup BBE Conf file C Program Files CCDC GOLD Suite GOLD examples tutoriall gold conf Load Save Options 14cM J Get ligands from Goldmine Wizard Templates Proteins Hostname asphalt Define Binding Site Select Ligands Port 8765 Waters Ligand Flexibility Number of GA runs 10 Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD I Send ligands to GoldMine E Constraints Atom Typing Hostname asphalt Port 8765 rM SQLite j Filename pup Suite GoldMine examples tutorial2 Cox2 db 7 J Database Hast User Password IT PostgreSQL N ASRS l Dock set v Read dock sets Help R Run GOLD Run GOLD In The Background Finish Cancel e Enable the Send ligands to GoldMine check box An appropriate machine Hostname and a Port number should be provided Then select either SQlite or PostgreSQL as the databas
307. structures are of blood coagulation factor Xa complexed with two different ligands The figure below shows a superposition of several experimental determinations of the factor Xa binding site complexed with a variety of different ligands not shown Only a small part of the binding site is displayed While it is clear that parts of the binding site are rigid their positions hardly moving from one structure to the next other parts are more inclined to move In particular the residue at the top right hand corner of the plot GIn192 adopts a variety of positions according to which ligand is bound The GIn192 position highlighted in purple is taken from 1lpg that shown in orange is taken from 1fax The next figure was produced by superimposing 1lpg and 1fax It shows the 1fax binding site and the 1lpg ligand Gln192 is highlighted in orange It is immediately clear that the 1lpg ligand cannot be docked accurately into the 1fax binding site if Gln192 is not allowed to move since there is a severe steric clash between these two GOLD User Guide 185 e To see this more clearly you can open Hermes and read in the file 1fax_ilpg_super mol2 from the folder containing the tutorial files via File Open this is the superposition from which the above figure was generated Preparation of Input Files e The file 1fax_protein mol2 contains the binding site from 1fax It has been set up for docking in the normal way Parts of the protein remot
308. sualiser display area 3 6 5 Defining a Binding Site from a List of Atoms or Residues e Click on Define Binding Site from the list of Global Options given on the left of the GOLD Setup window e Switch on the button labelled List of atoms or residues A file which contains a list of protein atom numbers or residues must be specified Either enter the path and filename of the file or click on the button and use the file selection window to choose the file 3 GOLD Setup Bik Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Atom select an atom in the visualiser or enter an atom index Proteins Define Binding Site zn 2N262 View Select Ligands Waters Point select atoms to define a centroid or edit XYZ Ligand Flexibility Fitness amp Search Options GA Settings x 6 4223 y 0 8690 Zz 15 3360 View Output Options GoldMine Parallel GOLD One or more ligands Constraints Atom Typing 4 reference ligand from 14 List of atoms or residues Filename fram Files CCDC GOLD examples tutorialS cavity_atoms txt se view Select all atoms within 10 fA J Generate a cavity atoms file From the selection Refine Selection IV Detect cavity restrict atom selection to solvent accessible surface J Force all H bond donors acceptors to be treated as solvent accessible
309. t in the Hermes visualiser those residues that have at least one of their atoms included in the binding site definition When entering a new value in the Select all atoms within box it is necessary to hit the enter key before the visualiser will update to reflect the changes made e After visual inspection you may wish to manually refine the binding site definition To do this switch on the check box labelled Generate a cavity atoms file from the selection By enabling this option the binding site definition will automatically be expanded to include all atoms in the existing definition plus all the atoms of their associated residues To manually refine this selection click on the Refine Selection button to open the Refine Binding Site Selection dialogue All residues included in the binding site definition are listed Residues can then be added or removed from the selection by clicking on atoms in the Hermes visualiser e The cavity atom selection can be saved as a protein atom subset and viewed within Hermes To do this click on the Add Definition as a Selection button You can then highlight the atoms belonging to the subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area GOLD User Guide 15 3 6 4 Defining a Binding Site from a Reference Ligand e Click on Define Binding Site from the list of Global Options given on the left of the GOLD Setup win
310. t lines are SP3_SP3_ BOND SP3_SP2 BOND SP2 SP2 BOND and UNKNOWN BOND The syntax is of the form SP3 SP3 BOND A n For example SP3_SP3_ BOND 0 18750 3 0 3 1515926 e The overall contribution of intramolecular strain to the scoring function is scaled by the coefficient called INTRA_COEFFICIENT in the ChemScore file see Altering ChemScore Fitness Function Parameters the ChemScore File 7 4 7 Covalent Term e When covalent bonding is switched on see Setting Up Covalently Bound Ligands the ChemScore function is modified in the following ways The clash term see Clash Penalty and Internal Torsion Terms is reduced so that no clash is registered for 1 2 or 1 3 contacts around the link atoms in the protein and ligand Torsion terms see Clash Penalty and Internal Torsion Terms are added for the rotatable parts of the linkage Avalence angle bending term is added to the overall energy to penalize poor link geometries e The weight of the covalent link energy in the ChemScore function is controlled by the parameter called LINK BEND COEFFICIENT in the ChemScore parameter file see Altering ChemScore Fitness Function Parameters the ChemScore File 7 4 8 Constraint Terms e Constraints see Setting Constraints are implemented in ChemScore in the same way as they are in GoldScore 7 4 9 Altering ChemScore Fitness Function Parameters the ChemScore File e The ChemScore param
311. t of output However it is possible to cut this down by applying output filter options These options can be used to Specify that all docking solutions are saved Retain only the n best docking solutions Save the top ranked solution for the best m ligands only GOLD User Guide 151 Filter out all solutions with fitness scores lower than a specified value e By default the Keep all solutions option from the Selecting Solutions panel in the Output Options window will be selected Starting the Docking Run e Weare now finished setting up our docking so click on the Run GOLD button at the bottom of the GOLD interface e You will be presented with a Finish GOLD Configuration window containing three Save Files options A Finish GOLD Configuration Directory C Users henderson Desktop tutorial 1 Save Files v GOLD conf file gold conf At least one protein has been edited 1ACM_protein mol2 V Protein s Save Cancel GOLD conf file if the gold conf has changed in any way or if there is currently no gold conf for the docking as is the case with this tutorial you will be provided with the option of saving out a gold conf file and or modifying its name Protein s in this case we have started from a raw PDB file that was not correctly set up for use with GOLD The modifications that we have made note we have been prompted that At least one protein has been edited mean that
312. tail below Further information on the content of all these output files is available see Description of Output Files The Ligand Log File gold_ligand_m1 log 154 Ten docking runs were set up for this ligand and for each of these docking runs the progress of the genetic algorithm is displayed in the gold_ligand_m1 log file displayed in the Run GOLD window The ligand log file is also saved to the specified output directory where m1 is the index to the number of the ligand in the input file Inspect the gold_ligand_m1 log in the Run GOLD window If you have closed this window by accident you can read the file from your output directory into a text editor and view it this way Following the completion of all docking runs on the ligand the results from the different runs are compared The end of the go ld_ligand_m1 log file will include a matrix of root mean square deviations RMSD between the various docked ligand positions see Comparison of Docking Solutions A clustering report is also given which can be used to identify different binding modes see Identification of Different Binding Modes Clustering of Ligand Poses It is possible that fewer than the specified ten dockings were completed due to the Allow early termination option being selected see Early Termination In the example output shown below the solution found for docking attempt number 2 has the best fitness score while the solution found for docking attempt number
313. tched can be found in the gold_ligand log file 83 8 9 84 Fragment file rotatable bond_override mol2 acylurea fragment no matches thioacylthiourea fragment no matches diarylamine fragment gt C ar NH C ar lt 1 matches Ligand bond 18 14 set to 1 Ligand bond 14 9 set to 1 sec amine 1 fragment 1 matches Ligand bond 14 9 set to 1 Ligand bond 14 18 set to 1 Ligand bond 14 15 set to 1 sec amine 2 fragment no matches sec amine 3 fragment no matches If using the postprocess instruction and rotatable bond override file the geometry is overruled whether the associated fitness flag is on or off If a torsion distribution can be found and matched this will be used to bias the geometry of the re typed bond Care should be taken to ensure the correct substructure is defined in the rotatable_bonds_override mol2 file If a substructure cannot be matched the bond override will not be used Fixing Rotatable Bonds at Their Input Conformation GOLD was designed to dock flexible ligands into protein binding sites However sometimes it can be useful to fix the geometry of part or all of the ligand e g in order to study the possible binding of a pre determined ligand geometry To fix rotatable bonds at their input conformation click on Ligand Flexibility from the list of Global Options given on the left of the GOLD Setup window and switch on the Fix Ligand Rotatable Bonds check box The following options are then available
314. ted with a Finish GOLD Configuration window containing Save Files options Finish GOLD Configuration Directory C Users henderson Desktop ensemble_docking output Save Files v GOLD conf file gold conf At least one protein has been edited 1TBF_protein mol2 1t9s_protein mol2 1xp0_protein mol2 2chm_protein mol2 v Protein s Cavity atoms cavity atoms Cancel e Ensure the GOLD conf file and Protein s tickboxes are activated and that the filenames are as you want them then hit Save to start the docking e As the job progresses output will be displayed in several tabs in the Run GOLD window GOLD User Guide 217 8 Run GOLD ESA list of ligand logs gold log gold_protein log gold err Messages ligand log This list will be updated every 10 seconds Click on a log to view it Interrupt GA View Solutions Close Once the job is complete load the docking results into Hermes by clicking on the View Solutions button in the Run GOLD window We have finished with the Run GOLD window now so close the window by clicking on the Close button In the GOLD Setup window click on the Cancel button to close this window as it is also no longer needed Viewing Results 218 Return to the Hermes 3D view and look at the Docking Solutions tab Let us think about what results we can expect We have loaded four proteins and one ligand Starting from a superimposed set of protein str
315. tein ligand complexes i e must be set up individually for each protein ligand if performing ensemble docking GOLD User Guide 91 10 2 1 92 Any distance between a ligand and protein atom or between two ligand or two protein atoms can be constrained to lie between minimum and maximum distance bounds GOLD features two types of distance constraint A standard distance constraint for use with individual ligands see Setting Up a Distance Constraint Asubstructure based distance constraint for use with multiple ligands which have a common functional group see Setting Up Substructure Based Distance Constraints Setting Up a Distance Constraint A distance between a specified ligand and protein atom or between two ligand or protein atoms can be constrained to lie between minimum and maximum distance bounds During a GOLD run if a constrained distance is found to lie outside its bounds a spring energy term is used to reduce the fitness score i e E kx where X is the difference between the distance and the closest constraint bound k is a user defined spring constant To set up a distance constraint you must first select the appropriate protein tab adjacent to the Global Options tab Select Distance from the list of Global Options given on the left of the GOLD Setup window If this option is not visible click on the icon next to Constraints to expand the list of options Specify the atoms to be used in t
316. term usually a weight in the range of 5 to 10 will work well GOLD User Guide gt GOLD Setup Of x Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial4 gold conf Load Save Options Protein 1cil lyase oxo acid Protonation amp Tautomers Waters Protein atom number 2041 Delete Ligands Flexible Sidechains Substructure file fingsthenderson Desktop gold_tutorialsjtutorial4 substructure mol2 Soft Potentials Metals 5 Constraints Substructure atom no 4 Distance P Use ring centre nearest to selected atom ring atoms only Substructure HBond Minimum separation 15 Protein HBond Covalent Maximum separation 3 5 Interaction Motif Spring constant 5 0 Reset Substructure protein 2041 C Documents and Settings henderson Desktop gold_tutorials tutor Help R Run GOLD Run GOLD In The Background Finish Cancel It is possible to define a distance constraint from a centroid of a ring in the ligand To do this specify an atom within the ring of interest and enable the Use ring center nearest to selected atom ring atoms only check box The closest ring center to the selected atom will be used Note When defining a distance constraint involving a ring center ensure that the maximum and minimum separations are adjusted accordingly If the constraint refers to a substructure atom and therefore a ligand atom which is topologically equivalent to other ato
317. ters Because we have defined the same value for RMSD as with our diverse solutions settings the cluster shortcuts will GOLD User Guide point to the top ranked solution in each of our diverse clusters see Identification of Different Binding Modes Clustering of Ligand Poses e Now hit the Run GOLD button to start the docking Change the GOLD conf file name e g to diverse_solutions conf this will ensure a new conf file is saved rather than overwriting the original file then hit Save to start the docking e Once the docking has completed load the results into Hermes by hitting the View Solutions button e Before closing the Run GOLD window inspect the go d_ligand_m1 log file Diverse solution information is given for each docked ligand under the heading Diverse Solutions Stats Move attempts 147731 Move failures 5705 Failure rate 0 039 e These stats are explained elsewhere see Method Used to Generate Diverse Solutions e Cluster information can be found at the end of the file e Atthe 2 44 Angstrom cut off there are 4 clusters e Close the Run GOLD window by hitting the Close button As before load the reference file ligand mol2 via File Open so that the docked poses can be compared to the crystallographic pose e You should see something similar to the above H atoms removed for clarity native ligand coloured by atom type Two of the solutions are close to the native binding pose GOLD User Guide 201 e
318. the Hermes visualiser e Select which ligands you wish to remove by switching on their corresponding Extract and Reload check boxes then click on the Extract button 8 GOLD User Guide Extracted ligands are removed from the protein file and automatically reloaded into Hermes so that they can be used e g to define the binding site see Defining a Binding Site from a Reference Ligand When extracting ligands you will be asked if you want to write the ligand to a file This can be useful for later comparison with docking results If the protein contains metal ions then GOLD can automatically determine their coordination geometry Virtual coordination points are then added at locations where GOLD is missing a coordination site and these coordination points are used as fitting points that can bind to acceptors see Metal lons However if you wish to delete a metal ion from the protein select the appropriate protein tab then click on Metals from the list of available options A list of the metal ions present in the protein file will be shown To remove a metal select it in the list and click on the Delete button Note that this is an expert GOLD option thus will be greyed out if you are using the wizard To make use of this feature you will need to exit the wizard 3 5 Water Molecules 3 5 1 Methodology For Handling Waters GOLD User Guide Water molecules often play key roles in protein ligand recognition Water molecules can either form m
319. the number of operators that are applied over the course of a GA run It is the key parameter in determining how long a GOLD run will take Changes to genetic algorithm parameters should be made with care see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 23 4 Number of Islands 226 Rather than maintaining a single population the genetic algorithm can maintain a number of populations that are arranged as a ring of islands Specifically the algorithm maintains n_islands populations each of size popsize GOLD User Guide Individuals can migrate between adjacent islands using the migration operator The effect of n_islands on the efficiency of the genetic algorithm is uncertain Changes to genetic algorithm parameters should be made with care see Controlling Accuracy and Speed with Genetic Algorithm Parameter Settings 23 5 Niche Size Niching is a common technique used in genetic algorithms to preserve diversity within the population In GOLD two individuals share the same niche if the rmsd between the coordinates of their donor and acceptor atoms is less than 1 0 A When adding a new individual to the population a count is made of the number of individuals in the population that inhabit the same niche as the new chromosome If there are more than NicheSize individuals in the niche then the new individual replaces the worst member of the niche rather than the worst member of the total population
320. this example we are going to use a set of coordinates to define the binding site The coordinates can either be input as x y z values or the centroid of a user selected group of atoms can be used We will use the former Activate the radio button that reads Point select atoms to define a centroid or edit XYZ The centre of the binding site in lacm can be found at 42 409 29 242 16 869 so enter these coordinates into the corresponding x y z boxes Click the View button in order to visualise those atoms included in the binding site definition The approximate radius of the binding site must also be specified By default the binding site radius is set to 10 0 A ensure that this is the case This radius should be large enough to contain any possible binding mode of the N phosphonacetyl L aspartate ligand A cavity detection algorithm LIGSITE is used to restrict the region of interest to concave solvent accessible surfaces Ensure that cavity detection is enabled by activating the Detect Cavity restrict atom selection to solvent accessible surface tickbox GOLD User Guide Click Next to proceed to the Configuration template dialogue Specifying a Configuration File Template Specifying the At this point you are given the option to load a configuration file template Configuration templates can be used to load recommended settings for a number of different types of docking protocols see Using Configuration File Templates In this e
321. ting Load Existing from the resultant pop up window and navigating to the directory where the gold conf is stored and clicking Open Distance Constraints GOLD User Guide Any distance between a ligand atom and a protein atom can be constrained or restrained to lie between minimum and maximum distance bounds GOLD features two types of distance constraint 173 A standard distance constraint for use with individual ligands see Standard Distance Constraints A substructure based distance constraint for use with multiple ligands which have a common functional group see Substructure Based Distance Constraints Standard Distance Constraints e Distance based constraints are specific to each protein thus click on the Protein 1cil lyase oxo acid tab to access all protein specific aspects of the docking setup e Hit the triangle adjacent to Constraints in the list of available options to expand the Constraints tree then select Distance 3 GOLD Setup _ oix Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial gold conf Load Save Options Protein 1cil lyase oxo acid Protonation amp Tautomers Waters Constrain distance from sah aa Minimum separation 15 en Maximum separation 3 5 vated Spring constant so Protein HBond I Use topologically equivalent atoms Covalent Interaction Motif Help R Run GOLD Run GOLD In The Background F
322. ting Up Substructure Based Distance Constraints HBond constraint see Setting Up Hydrogen Bond Constraints Protein HBond constraint see Setting up Protein H Bond Constraints Interaction motif see Setting up an Interaction Motif Constraint Note Protein specific constraints are only evaluated if the respective protein structures are selected for scoring in the ensemble docking process It is also possible to only specify a constraint for a single protein structure in the ensemble There are caveats associated with definition of constraints when docking into an ensemble see Caveats of Docking into Ensembles It is not possible to apply soft potentials or covalent constraints to a docking ensemble Interpreting Ensemble Docking Output Standard docking output is detailed elsewhere see Viewing and Analysing Results The following details ensemble specific output Each initialised protein is written to a file of the type gold protein lt ensemble index gt mol2 Each solution file will contain a gt lt Gold Ensemble ID gt tag with the ensemble index identifying the protein that GOLD has selected as the receptor for this solution gt lt Gold Goldscore Internal Correction Weighted gt 1 8964 gt lt Gold Ensemble ID gt The ligand rnk file and the bestranking 1st file have an additional ensemble column that details the ensemble index of the protein the ligand was docked into The gold_protein 1log wi
323. to create it with the GOLD graphical front end the file can be written out when the Run GOLD Run GOLD in the background amp Finish buttons are hit A number of configuration file templates are also available see Using Configuration File Templates In addition GOLD uses a parameter file see Altering GOLD Parameters the gold params File a scoring function specific parameters file see Fitness Functions and optionally a torsion distribution file see Using Torsion Angle Distributions All these files are supplied in the GOLD distribution and by default will be found automatically by the program If required any of the files can be copied to a user s directory and edited and GOLD can then be directed to use the edited file 14 2 Running GOLD Interactively 122 GOLD can be run interactively by hitting the Run GOLD button in the GOLD Setup window Before the job is started you will prompted to save a configuration file The configuration file is a text file which specifies the GOLD calculation that is to be run including details of the ligand the protein binding site the fitness function parameter file to be used the torsion distribution file to be used and the genetic algorithm parameters see Saving and Re using Program Settings in Configuration Files To save a configuration file specify the Directory or click on the button and use the browse for folder window to choose the directory Then enable the GOLD conf
324. tor H A distance R_IDEAL 1 85 in A Ar The absolute deviation of the actual H A Calculated for each H bond separation from r AF ideal The tolerance window around the H A DELTA_R_IDEAL 0 25 distance r within which the H bond is regarded as ideal GOLD User Guide 63 Term AF max Or Term a Aa A Q ideal A QA max Oa D H A distance parameters D Donor A Acceptor Meaning Name in ChemScore File Default Value The maximum possible deviation from the DELTA_R_MAX 0 65 ideal distance above this the interaction is not regarded as an H bond The Gaussian smearing sigma associated with HBOND_R_SIGMA 0 1 this term D H A angle parameters D Donor A Acceptor Meaning Name in ChemScore File Default Value The ideal D H A angle in degrees ALPHA IDEAL 180 0 The absolute deviation of the actual D H A Calculated for each H bond angle from The tolerance window around the D H A DELTA ALPHA IDEAL 30 0 angle within which the H bond is regarded as ideal The maximum possible deviation from the DELTA ALPHA MAX 80 0 ideal D H A angle above this the interaction is not regarded as an H bond The Gaussian smearing sigma associated with HBOND ALPHA SIGMA 10 0 this term DH A X acceptor centred angle parameters D Donor A Acceptor X Heavy atom attached to A Term p Ap A P ideal A Pma Op 64 M
325. tors are treated as part of the protein Atom atom potentials were calculated for each atom pair with an excess of 150 observations in the database using the ASP reference state for atom types with fewer observation the potential was set to zero for all distances For short distances there will be no observed contacts and the potential is set to 10 The atom atom potentials for all atom types can be found in the GOLD installation directory GOLD_HOME gold asp tables The statistical potentials are augmented with the ChemScore clash term and internal energy term see Clash Penalty and Internal Torsion Terms The internal energy GOLD User Guide term is needed to prevent the docking of high energy ligand conformations while the clash term should prevent protein ligand clashes where the supplied potential is too soft to provide sufficient repulsion between protein and ligand atoms and at the same time preventing overlap between atoms with no potential i e too few observations for the generation of a non zero potential The final ASP fitness can be written ASP Fitness C yy StatScore p l rod _ Cite int Clash ay int Ctasn clash EE StatSeoret l rp S map p e The total StatScore written as S map in the ligand output file is a summation over all combinations of protein atoms p and ligand atoms within 6 0 A and rp is the distance between protein atom p and ligand atom C is a scaling factor and Cin and Cclash are
326. traints in the Protein 1qbt aspartyl protease tab and select Protein HBond from the list of constraint types 165 GOLD Setup BBE Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial3 gold conf Load Save Options Protein igbt aspartyl protease Waters Delete Ligands Flexible Sidech Soft Potentials Metals Constraints Distance HBond Covalent Protonation amp Tautomers Substructure Protein HB Interaction Motif Protein atomis required to Form H bond ains Constraint weight 10 0 Minimum H bond geometry weight 0 005 ond Run GOLD Run GOLD In The Background Finish Cancel When specifying a protein hydrogen bond constraint the protein atom must be selected in the 3D view GOLD will then be biased towards finding solutions in which the specified protein atom forms hydrogen bonds However as with standard hydrogen bond constraints such a solution is not guaranteed During the GOLD run the fitness score of a given docking will be penalised for every protein H bond constraint that is not satisfied The Constraint weight is the strength of bias applied to the formation of a specified hydrogen bond in the least squares mapping algorithm within GOLD The Constraint weight is also the value of the penalty applied to the fitness score for each constrained H bond that is not formed The Minimum H bond geometry w
327. trogen amide N am Nitrogen trigonal planar e g in nitro pyrrole N pl3 Nitrogen sp3 positively charged e g in lysine N 4 Oxygen sp3 0 3 Oxygen sp2 0 2 Oxygen in carboxylates and phosphates O co2 Sulphur sp3 S 3 Sulphur sp2 S 2 Sulphoxide sulphur S o Sulphone sulphur S 02 Phosphorus sp3 P 3 Halogens metals normal element symbols e g F Cl Ca Zn Bond types Single 1 Double 2 Triple 3 Aromatic ar Amide am delocalised e g in carboxylate guanidinium ar 220 GOLD User Guide 22 Appendix C Additional Tags in Output Files e Solution output files for the docked ligand s can contain additional information such as the scoring function terms and the rotated protein hydrogen atom positions that were generated during the docking e This information can be written to SD file tags for MOL2 files these tags are written to comment blocks This additional information is particularly important when post processing docking results with GoldMine It is possible to control the information written to solution files see Controlling the Information Written to Ligand Solution Files e The table below lists the tag names that you are likely to see in GOLD solution files Name Gold Protein ActiveResidues Gold Protein RotatedAtoms Gold Protein RotatedWaterAtoms Gold Protein RotatedTorsions Gold Id Protein Gold Id Ligand Gold Rescore Rmsd Scoring terms Gold Fitness Score Gold Covalent Energy Gold Constraint Score
328. ts list K GOLD Setup l 7 EE Conf file C Users henderson Desktop tutorial3 gold conf Load Save Global Options Protein igbt aspartyl protease Protonation amp Tautomers Extract Delete Waters Protein atom s required to form H bond 242 241 Delete Ligands Flexible Sidechains SEES 10 0 Soft Potentials Minimum H bond geometry weight 0 005 Metals 4 Constraints Distance Substructure HBond Protein HBond Covalent Interaction Motif Reset Type Details Protein HBond 10 0 0 005 242 241 Help R Run GOLD Run GOLD In The Background Finish Cancel e Specify protein H bond constraints for the three remaining key hydrogen bonding interactions as outlined in the table below note that you may have to hit the Reset button to clear the Protein atom s required to form H bond window Note It is necessary to specify the hydrogen atom to define the donor partner in the H bond constraint ia pibending Atom number s Constraint weight Sey ii Ile50 chain A 1914 10 0 0 005 Ile50 chain B 2724 10 0 0 005 Asp25 chain B 1161 or 1162 10 0 0 005 e Once all of these protein H bond constraints have been set up the Constraints Editor window should contain four individual constraints 168 GOLD User Guide GOLD Setup BBE Conf file C Documents and Settings henderson Desktop gold_tutorials tutorial3 gold conf Load Save Options Protein igbt aspartyl protease Waters Metals
329. uctList aspx The ligand is an essential co factor in enzymatic transamination and is bound covalently through an azomethine linkage between the ligand and an active site lysine LYS258 This tutorial will illustrate how to carry out a covalent docking by docking the PLP N oxide ligand back into its native binding site Preparation of Input files The original PDB file 1ASE pdb has been provided should you wish to set up the protein and ligand files yourself Protein and ligand files are also provided and have been set up in accordance with guidelines for the preparation of input files Setting Up the Protein s and Setting Up Ligands respectively Running a Covalent Docking GOLD makes a few assumptions when docking covalently see Method Used for Docking Covalently Bound Ligands It is assumed that there is just one atom linking the ligand to the protein The link atom must be present in both the protein and ligand files Ideally in both files the link atom will have a free valence available through which the link can be made It is possible to dock covalently to a single ligand see Setting Up a Single Covalent Link or a ligand substructure see Setting Up Substructure Based Covalent Links In this case we are docking only to a single ligand Note that mol2 files must be used when running covalent dockings Load both the protein and ligand files into Hermes You will see that the ligand is indeed covalently bonded to LYS
330. uctures GOLD evolves a separate population of individuals representing ligand conformations for each protein structure that is part of the ensemble The best ligand conformation found in any of the ensemble structures is returned i e GOLD selects the best protein for a particular ligand based on the max fitness value of a ligand For example if for a given GA run a ligand gets the scores 10 in protein 1 20 in protein 2 and 15 in protein 3 protein 2 will be selected Return to the Hermes 3D view and inspect the top ranked solution predicted by GOLD Note that the original four proteins are still loaded To make the display less complicated you may wish to disable these four proteins by deactivating the tickbox adjacent to 1t9s 1TBF 1xp0 and 2chm under the Display tab in the Molecule Explorer f you do this return to the Docking Solutions tab once you have finished The docking solutions are given in their docked order with their corresponding fitness score listed under the column headed PLP Fitness If required the solutions can be ordered by clicking on this PLP Fitness header to determine which is the highest scoring GOLD User Guide We have obtained 20 docking solutions as this is how many times our ligand was docked The protein the solution corresponds to may be one of four identifiable by the ensemble index number 1 4 The initialised protein is given a filename of the type gold protein lt ensemble index gt mol2 Loosely
331. ults Alternatively GOLD can be run again following the instructions below e Load the gold conf into GOLD via GOLD Setup and Run a Docking Load Existing then navigate to the folder containing the tutoria18 files select gold conf and click Open This automatically loads the settings and parameter values for this tutorial into the GOLD front end e No settings need to be changed for the purposes of this docking however you may wish to change the output directory To do this click on Output Options and either type the path to or browse to an appropriate output folder where the GOLD output files can be saved e The GOLD run is started by hitting the Run GOLD button at the bottom of the interface In the Finish GOLD Configuration window you will be prompted that the GOLD configuration has been updated and needs to be saved We have not modified the protein mol2 file so we do not need to save this file thus ensure the tick box adjacent to protein mol2 is deactivated Change the configuration file name then hit Save to start the GOLD run e Once the docking is complete the message Finished Docking Ligand ligand mol2 will appear in the gold_ligand_m1 log tabbed view of the Run GOLD window The GOLD results can be read into Hermes via the View Solutions button in the Run GOLD window e The input ligand can also be used as a reference ligand i e the original pose of the ligand in the crystal structure so read this in via File Open e The 3D
332. un has finished hit Close in the Run GOLD window Running GOLD Dockings All waters turned off Return to the GOLD front end and click on Configure Waters to bring up the water setup window Change the toggle state of each water molecule to off Go to Output Options and change the output sub directory name e g to waters_off Hit the Run GOLD button In the Finish GOLD Configuration window as before edit the name of the GOLD configuration file in the GOLD conf file text box to e g waters_off conf Hit Save There is no need to change any other settings Once GOLD has finished hit Close in the Run GOLD window then as we have finished with GOLD hit Cancel in the GOLD setup window Tidy up the 3D view by going to File then selecting Close All Files Analysis of results All waters turned on 182 Load the results of the waters on docking by hitting File then Load GOLD results and navigating to where waters_on conf is stored Now read the reference ligand by clicking on File Open then navigate to the tutorial 5 directory which contains the ligand_reference mol2 file Select the file then click Open Scroll through the docking solutions one by one to check their poses against that of the reference ligand You should find that several docking solutions are found none of which closely resemble the correct binding mode Note You may wish to modify the representation style of the reference ligand so that it is more clear Hit the
333. und between the solutions and rank file Increasing the number of dockings or the number of GA operations in each docking will result in the discrepancy being less pronounced 9 2 Hydrophobic Fitting Points 86 GOLD automatically calculates a list of hydrophobic fitting points in the binding site These are used during the generation of trial docking solutions to map hydrophobic ligand atoms into favourable regions of the binding site GOLD generates its hydrophobic fitting points by placing a fine grid over the binding site At each grid position the van der Waals interaction energy between a bare carbon atom and the protein is evaluated By default positions at which the interaction energy is below 2 5 kcal mol are added to the list of fitting points The potential and threshold for selecting fitting points can be changed by editing the gold params file and changing the values of INTERNAL POTENTIAL FITPTS and E FITPT THRESHOLD In this way a map is constructed that contains positions onto which the placement of a hydrophobic ligand atom should be favourable The ligand fitting points are used for the matching of hydrophobic regions By default only carbon atoms in the ligand are considered when identifying fitting points The selection of suitable ligand atoms can be extended to include carbon halogen and non polar sulfur atoms by uncommenting the following line in the gold params file LIGAND FITPTS SELECTION EXTEN
334. up i yx Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial2 gold conf Load Save Options Protein 1442 lyase Wizard Templates Ligand File GA Runs First Ligand Last Ligand Proteins x z5 Define Binding Site 1 ligand mol2 10 1 last Select Ligands Waters Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing Click on the Add button and use the file selection window to choose the ligand data file s then specify the number of times each ligand is to be docked by entering a value in the GA runs box J Show full file paths Add Delete Reference ligand Bas Help h Run GOLD Run GOLD In The Background Finish Cancel e Balloon help can also be viewed by right clicking at certain locations in the interface and selecting What s this from the resulting menu GOLD User Guide 139 18 References GOLD references Molecular Recognition of Receptor Sites Using a Genetic Algorithm with a Description of Desolvation G Jones P Willett and R C Glen J Mol Biol 245 43 53 1995 Development and Validation of a Genetic Algorithm for Flexible Docking G Jones P Willett R C Glen A R Leach and R Taylor J Mol Biol 267 727 748 1997 Protein Ligand Docking and Virtual Screening with GOLD J C Cole J W M Nissink R Taylor in Virtual Screening in Drug Discovery Eds
335. used here is acetylcholine esterase PDB entry code 1acj the protein that more than any other is essential for the correct transmittal of nerve impulses in the brain and around the body The ligand is tacrine an inhibitor of acetylcholine esterase which is a drug used to treat Alzheimer s disease The active site of the enzyme has been modelled with three water molecules in it each of which makes hydrogen bonds with the protein This tutorial will illustrate the requirements for setting up and running dockings in which the protein binding site features one or more water molecules The example chosen mimics the situation where a researcher has a crystal structure of a protein binding site and is unsure which and how many of the waters in that binding site should be included in the model for use in an inhibitor design effort Preparation of Input Files GOLD User Guide Open Hermes and read in the file protein mo 2 from the folder to which you copied the tutorial5 files The acetylcholine esterase protein mol2 has already been set up in accordance with the guidelines for the preparation of protein input files see Setting Up the Protein s The full protein is not displayed Parts of the protein remote from the binding site have been deleted in order to speed up the calculation see Essential Steps Hydrogen atoms have been placed on the protein in order to ensure that the ionisation and tautomeric states are defined unambiguously see
336. ustering with the complete linkage method 2 3 4 5 6 7 8 9 1 0 8 1 1 1 0 1 0 1 4 2 3 5 0 4 6 2 0 9 1 1 1 1 1 2 2 3 5 2 4 6 3 0 4 0 8 0 9 2 3 5 0 4 5 4 0 6 1 1 2 3 4 9 4 5 5 1 3 2 0 4 9 4 5 6 1 8 5 1 4 4 229 7 5 3 4 5 8 2 4 Step Distance between clusters being merged Clusters 1 0 40 1 2 3 4 9 5 6 71 8 2 0 84 1 2 3 4 5 9 8 6 3 0 84 1 2 7 3 4 5 91816 4 1 13 1 2 3 4 5 7161918 5 1 42 1 2 3 4 5 6 71819 6 2 35 1 2 3 4 5 6 7 918 7 2 38 1 2 3 4 5 6 7 8 9 8 5 28 1 2 3 4 5 6 7 8 9 24 3 identify_ligand py e Locatedinc Program Files x86 CCDC goldsuite 5 3 GOLD gold d_win32 bin on Windows machines e identify_ligand py can be used to extract a specific ligand description from PDB SDFile or MOL2 format input files e t requires a filename and a ligand number n as arguments and then locates the nth ligand in the file If any descriptive information such as the ligand name is available for that ligand it is then displayed e identify_ligand py can be invoked from the command line The structure of the command is identify_ligand py lt ligand data file gt lt ligand number gt Note identify_ligand py is a Python script and as such requires a working installation of Python http www python org 24 4 check_mol2 exe e Locatedinc Program Files x86 CCDC goldsuite 5 3 GOLD gold d_win32 bin on Windows machines e check _mol2 uses the same algorithms as the main GOLD program to ch
337. utions ccccsesseecceceeeeeesseeeceeesesaaeeeeeeeess 233 25 3 Example Torsion Angle Distributions c cccsseseeeceececeeeeeseeeeeeeessaaaaseeeeeess 235 25 4 Extracting Torsion Angle Distributions from the Cambridge Structural Database GOLD User Guide 1 Introduction GOLD User Guide GOLD Genetic Optimisation for Ligand Docking is a genetic algorithm for docking flexible ligands into protein binding sites GOLD is supplied as part of the GOLD Suite which includes two additional software component Hermes and GoldMine The Hermes visualiser can be used to assist the preparation of input files for docking with GOLD visualisation of docking results and calculation of descriptors The Hermes visualiser is also used for interactive docking setup e g for defining the binding site and the setting of constraints GoldMine is a tool for the analysis and post processing of docking results see Overview of GoldMine Further details are provided in the Hermes and GoldMine documentation GOLD provides all the functionality required for docking ligands into protein binding sites from prepared input files see Setting Up the Protein s and Setting Up Ligands Although Hermes can be used to assist the preparation of input files e g the addition of hydrogen atoms including those necessary for defining the correct ionisation and tautomeric states of protein residues GOLD will likely be used in conjunction with a mode
338. utions Click on Output Options from the list of Global Options given on the left of the GOLD Setup window and enable the Create links for different binding modes based on GOLD User Guide RMSD clustering check box and specify the Distance between clusters this determines how similar the poses are in each cluster of solutions By default the clustering distance is 0 75 A e Aclustering report is given at the end of the ligand log file see Ligand Log File The clusters themselves and the individual solutions within each cluster are in ranked order i e the first member of the first cluster is always the top ranked solution For example output from a run of 10 GA dockings may look like Final ranked order of GA solutions 8 3 610 7 45 9 2 1 RMSD Matrix of RANKED solutions 2 3 4 5 6 T 8 9 10 1 0 6 0 6 0 8 0 5 6 2 0 8 1 1 5 9 6 1 22 Ow Dd Oa 6 4 079 222 621 623 3s 0 7 0 4 6 1 0 7 1 0 5 9 6 1 4 0 6 5 8 0 4 1 0 5 6 5 8 Da 6 1 0 6 1 0 5 9 6 1 6 5 9 5 7 0 9 0 6 fos 120 5 7 Dg es E Dick 9 0 8 Clustering method complete linkage Structure ids in cluster table rank nos Ordering of clusters and their members by rank Corder if from rms_analysis Distance Clusters 0 37 Ty 2 38 Bel 4 6 Be SP 20 0 39 A 2b Be AF eed eT 9 lO 0 58 13 5 2 4 7 6 8 9 10 0 59 13 5 2 4 7 610 8 9 0 72 123 5 4 7 620 8 9 0 90 a 3 Se Lf be 29 0 8 lt files d 0 75 A 0 96 12 3 5
339. ve extracted the important waters all other waters must be deleted from the protein file This is done by hitting the Delete Remaining Waters button Ifthe waters are extracted in this way they are automatically added to the Configure Waters dialogue under the Global Options tab e By default each water molecule in the Configure Waters list will be retained in the binding site during docking and will be allowed to spin in order to optimise the orientation of the water hydrogen atoms These settings can be customised for specific water molecules within this dialogue 3 GOLD Setup Loix Conf file C Program Files CCDC GOLD examplesitutorial5 gold conf Load Save Options Protein 1acj hydrolase carboxylic esterase Wizard Templates index toggle state spin st Proteins Define Binding Site Protein 1acj hydrolase carboxylic esterase _HOH150 3 toggle ffspin Select Ligands Waters toggle 7 spin Ligand Flexibility Fitness amp Search Options Protein 1acj hydrolase carboxylic esterase _HOH152 3 GA Settings Output Options GoldMine Parallel GOLD Constraints Atom Typing Add Delete Help h Run GOLD Run GOLD In The Background Finish Cancel e For each water molecule listed the following can be specified The state of the water available options are On use the water for docking i e present Off do not use the water for docking i e absent Toggle have GOLD decide wheth
340. ve modified any of the rotamer settings from those initially loaded hit Rigid to reset the rotamer settings then hit Library again Hit Accept to close the rotamer definition window e GLN192 is now listed as being constrained to nine rotamers in the Flexible Sidechains window e We only require GLN192 to be flexible for the purposes of this example however using this method we can specify up to 10 rotatable side chains if we wished e Click on the Global Options tab and within the Output Options window change the output directory to e g flexible2 then hit the Run GOLD button e Change the name of the gold conf to e g flexible2 conf then hit Save Once the docking has finished load the results into Hermes using the View Solutions button e The following describes the output in the flexible directory provided with the tutorial If you have set up and run your own flexible docking using the instructions above your output may vary slightly however the general trends should be the same e Compare the top ranked solution with the experimental position of the 1lpg ligand The top ranked solution from the flexible run is much better It is not perfect in particular the benzamidine moiety is somewhat displaced but the benzyloxy side chain is now roughly in the right position the GIn192 side chain having moved out the way reference ligand C atoms coloured grey docking result C atoms coloured green GOLD User Guide 191 Also the
341. w A Fitting points summary is provided in the gold_protein log file The polar fitting points used by GOLD are also saved as protein atom subsets within Hermes Two subsets are saved donor hydrogens and lone pairs You can highlight the atoms belonging to any subset by picking the required subset from the Atom Selections pull down menu which is situated above the visualiser display area 19 3 7 Rotatable O H and NH Groups The torsion angles of Ser Thr and Tyr hydroxyl groups will be optimised by GOLD so their starting positions do not matter Specifically each Ser Thr and Tyr OH will be allowed to rotate to optimise its hydrogen bonding to the ligand Lysine NH3 groups are similarly optimised unless they are held in place by strong H bonds to neighbouring protein residues The optimised positions of polar protein hydrogen atoms generated during docking can be written to GOLD solution files see Controlling the Information Written to Ligand Solution Files It is possible to run a docking keeping these rotatable bonds static if required see Docking into a Rigid Protein 3 8 Docking into a Rigid Protein Even when not using advanced protein flexibility see Protein Flexibility serine threonine and tyrosine hydroxyl groups are optimised i e rotated during docking as are lysine NH3 groups In some cases it might be necessary to dock into a rigid protein i e to keep all the polar hydrogen atoms fixed during a docking
342. w use of a faster scoring function for docking The validation experiments will be reported on the CCDC website and in the open literature Although these protocols are currently believed to be optimum they should be used with care as the datasets used to derive these protocols are small Other protocols may work better for individual target proteins It is recommended that ChemPLP be used for target classes not mentioned here To load a template configuration file click on Templates from the list of Global Options given on the left of the GOLD Setup window Select the template you wish to use from the list of available templates then click on the Load Template button Note that configuration file templates are independent of the protein and ligand input files so these will need to be specified in the usual way before running the docking 16 3 Customising Scoring Function Parameters Empirical parameters used in the fitness function hydrogen bond energies atom radii and polarisabilities torsion potentials hydrogen bond directionalities etc are taken from the GOLD parameter file These parameters are independent of the scoring function being used Parameters can be customised by copying the file editing the copy and instructing GOLD to use the edited file see Altering GOLD Parameters the gold params File A scoring function specific parameters file is also used for GoldScore this is called goldscore params Parameters within this f
343. window obtained by clicking on the button adjacent to the Protonation Rules window The file can be modified and supplemented to suit user preferences 3 3 3 Flipping Asn and Gin Residues Terminal CO NH groups in Asn and Gin residues can be flipped i e rotated 180 deg This can be useful when dealing with poorly resolved protein structures in which you suspect the oxygen and nitrogen atoms may have been incorrectly determined i e transposed As residues are protein specific click on the appropriate protein tab adjacent to the Global Options tab and select Protonation and Tautomers from the list of options provided A list of the Asn and Gln residues within the defined binding site will be displayed Select the residue you wish to flip from this list the selected residue will be highlighted in the Hermes visualiser and click on the Flip button in order to rotate the CO NH group 180 degrees 3 3 4 Specifying Histidine Tautomers GOLD User Guide GOLD will not vary tautomeric states during docking e To specify the tautomeric state of particular histidine residues within the binding site select the appropriate protein tab then select Protonation and Tautomers from the list of options given on the left of the GOLD Setup window e A list of the His residues within the defined binding site will be displayed Select the His residue you wish to edit from this list the selected residue will be highlighted in the Hermes visualis
344. xample we will specify all docking settings manually Click Next to proceed to the Select ligands step Ligand File As with the protein file all hydrogen atoms must be present in the ligand input file see Ligand Hydrogen Atoms lonisation States and Tautomeric States We have already added H atoms to our ligand extracted it from the protein binding site and saved it From within the Select Ligands window it is possible to Add single ligands Select a complete directory of ligand files Specify a single file containing several ligands i e a multi MOL2 or SD file Specify the ligand you saved earlier by hitting the Add button at the bottom of the GOLD Wizard Navigate to folder to which you copied the tutorial1 files select ligand mol2 then click Open The Ligand mo12 will be listed under Ligand File The number of dockings to be performed on each ligand is specified under GA runs by default this value is 10 The value can be edited by clicking in this window and re entering another value however 10 GA runs are sufficient for this docking Click Next to proceed to the Choose a fitness function window Selecting a Fitness Function GOLD User Guide During a docking run the solutions found by GOLD are scored according to a fitness function see Fitness Functions GOLD offers a choice of fitness functions Piecewise Linear Potential CHEMPLP see Piecewise Linear Potential CHEMPLP GoldScore see GoldScore ChemScore
345. y docking into subtly different versions of the same protein see Ensemble Docking by docking using soft potentials see Allowing For Localised Movements Docking With Soft Potentials 4 1 Side Chain Flexibility 4 1 1 Introduction You may specify that one or more protein side chains are to be treated as flexible Each flexible side chain will be allowed to undergo torsional rotation around one or more of its acyclic bonds during docking Making a side chain flexible can make docking more difficult because it increases the search space that must be explored It may also increase the chance of false positives i e ligands that appear to dock well but do not actually bind Therefore you should only make a side chain flexible if you have good reason to believe e g from X ray data that it is likely to move in response to ligand binding 4 1 2 Specifying Flexible Side Chains You may specify that one or more protein side chains are to be treated as flexible during docking Flexible side chains are protein specific thus click on the protein tab adjacent to the Global Options tab in the example below the protein tab is named Protein 1fax coagulation factor then select Flexible Sidechains from the list of available options A list of the side chains included within the binding site definition will be displayed GOLD User Guide 25 26 GOLD Setup Big ks Conf file C Program Files CCDC GOLD Suite GOLD examples tutorial
346. y termination option instructs GOLD to terminate docking runs on a given ligand as soon as a specified number of runs have given essentially the same answer In this situation it is probable that the answer is correct and GOLD will just be wasting time if it performs more docking runs on that ligand To switch early termination on click on Fitness and Search Options from the list of Global Options given on the left of the GOLD Setup window then enable the Allow early termination check box To specify the early termination criterion click on the Early Termination Options button In the example below GOLD has been instructed to stop docking a ligand if it reaches a state in which the best three solutions found so far are all within 1 5 A rmsd of each other 107 11 3 11 3 1 108 Conf file c Program Files CCDC GOLD Suite GOLD examples tutorial9 gold conf Load Save Options Protein 1ase aminotransferase Wizard Templates IV Docking Proteins Define Binding Site Scoring Function coldScore z Select Ligands waters Parameter file DEFAULT i Ligand Flexibility Fitness amp Search Options GA Settings Output Options GoldMine pT Rescore caaan Early Termination Options 21 xi Atom Typi Terminate the number of GA runs early if the top E solutions are within 1 5 A Options IV Allow early termination Early Termination Options Generate diverse solutions

User Guide - The Cambridge Crystallographic Data Centre

Contents

Download Pdf Manuals

Related Search

Related Contents