Home

IsoStar User Guide and Tutorials 2010 Release

1. 5 H gets a score of 3 H and H also get scores of 3 Scores based on summing the atomic numbers of bound atoms are also computed Cy gets a score of 13 Cpm gets a score of 8 and the protons all score 6 This means that Cy is distinct from Cy 4 e In the third cycle of iteration the scores based on numbers of bonds become 5 for all the protons but the scores based on atomic numbers become 13 for H and 8 for H and H Thus H is distinct from H and Hp e The termination criterion for the iterative process is when no further atoms can be assigned as unique by an iteration At this point we know which atoms are grouped together those that had the same score at each iteration are topologically equivalent In this example the fourth pass shows that H and H are equivalent TsoStar User Guide 77 78 Appendix 3 References e Isostar A library of information about non bonded interactions I J Bruno J C Cole J P M Lommerse R S Rowland R Taylor and M L Verdonk J Comput Aided Mol Des 11 525 537 1997 The Protein Data Bank H M Berman J Westbrook Z Feng G Gilliland T N Bhat H Weissig I N Shindyalov and P E Bourne Nucleic Acids Res 28 235 242 2000 TIsoStar User Guide Appendix 4 Acknowledgements The CCDC gratefully acknowledges e The members of the Research Collaboratory for Structural Bioinformatics RCSB responsible for the maintenance of the PDB e Roger Amos and Anthony Sto
2. e measurement of contact distances and angles see Section 5 8 e Theoretical energy minima see Section 4 1 and crystal statistics data see Section 4 2 are tabulated in web browser pages and the former can be displayed in the visualiser 1 3 Calculating New Scatterplots e An ancillary program IsoGen see IsoGen Quick Summary page 53 allows you to calculate scatterplots for interactions that are not present in the standard IsoStar library TsoStar User Guide 2 Organisation and Navigation of IsoStar Data 2 1 2 2 2 3 Central Groups Information in IsoStar is accessible via a web browser interface and is organised on a per group basis e g there is a page that provide access to information on the nonbonded contacts formed by the group NO a page that gives analogous information for CH CH etc In IsoStar these groups are called central groups see Section 1 1 There are over 300 central groups in the IsoStar library ranging from single atom substituents such as fluoro common linking groups such as ester ring systems such as isoxazole amino acid side chains and common solvate molecules Contact Groups For each central group see Section 2 1 information is held on its interactions with up to about fifty contact groups see Section 1 1 The same set of contact groups is used for all central groups They fall into the following categories e General groups viz any C N O S or H atom and any polar hydr
3. 1sogen lt search name gt 4 2 Basic Options in the IsoGen Interface e On starting IsoGen the user is presented with a graphical interface from which to produce scatterplots File View Expert Run Help exam ple cor A FF Coordinate file example istr Isostar file name Oe SSS Ff Central group atoms ledgds Central group name Mo Mame Given mE oo 1 Contact group atoms 9 Contact group name No Name Given it J Allow short contacts Switch groups m l e The IsoGen interface in its Basic mode contains a number of items the functions of which are summarised below e Coordinate file The name of the cor file produced by a ConQuest search e IsoStar file name see Section 4 2 1 58 TsoStar User Guide e Central group atoms see Section 4 2 2 e Contact group atoms see Section 4 2 3 e Central group name The name that will be used for the central group in the IsoStar graphical interface By default this 1s set to No Name Given e Contact group name The name that will be used for the contact group in the IsoStar graphical interface By default this 1s set to No Name Given e Those superscripted by a star are mandatory for successful completion of IsoGen but are in most cases read in automatically from the ConQuest output files Usually the defaults are adequate to produce an acceptable scatterplot In such cases all the user need do 1s click on Run in the top level menu
4. 96 Go to the contact group table and hit the round purple button in the row marked any polar X H and the column headed Stats This displays some simple statistics about N O S H carboxylate contacts in the CSD TsoStar User Guide Statistics for contacts between charged carboxylic acid and any polar X H X N O or S More Info on the Statistics Source Nb Ne 100 NcINb ee csp 5000 1661 S9 1 0 3 Subset from 5358 crystal structures Nb the number of crystal structures that have both the central and contact groups present Ne the number of crystal structures in which the central group and contact group form a contact with distance A less than the sum of van der Waals radii v drel relative density is defined as dre dshort diong where dsnort is the density of contact groups with A lt V and diong is the density of contact groups with VER lt VY 0 5 e The column headed 00 N_ N indicates that these contacts occur in a high percentage of structures implying that they are very favourable energetically The last column d o contains a simple statistic that shows how common short contacts are i e shorter than the sum of van der Waals radii compared to long contacts The higher the number the more frequently short contacts occur e In this case the number is very high a value of lt 1 is probably typical for most other contacts This suggests that short N O S H carb
5. DRAW EDIT ERASE ADD 3D Options CONTACT Jp By Delete Contacts 2 a C C 1 pra ai or C N H 10 Options Delete Contact Distance Range You must specify a distance range for Contacts involving centroids E is Modify options and hit OK when done Distance range RingMaker From lo to PE A Range 4 C Value tolerance Templates e We will define the distance to be the sum of van der Waals radii plus a tolerance of 1 0 A This gives a range of 0 to 4 5 A which should be entered in the two distance range boxes Click OK twice to close the Distance Range and Non bonded Contact Definition windows and then Done to close the Geometric Parameters window 4 Running the Nonbonded Search e Hit Search in the sketcher window A Search Setup window will pop up 106 TsoStar User Guide Filters Advanced Options Search Setup Search Name search Available Databases 3D coordinates determined l Rfactor lt 0 05 O lt 0075 M CSD version 5 30 November 2008 CM lt 0 1 Not disordered No errors You can search complete database s or a subset e g hits found in a previous search Select Subset single query being used Search will find structures T Not polymeric M Noions M No powder structures M Only Organics where this query is true Start Search e Click on the Advanced Options tab and enable the Normalise terminal H positi
6. TsoStar User Guide 69 5 5 70 variable This may be done with the Omit atoms option As the name suggests this option makes it possible for user specified central group atoms to be omitted from the final scatterplot Atoms which are omitted in this way are not displayed in the scatterplot not used in superimposing the central group fragments or calculating the average central group geometry and not used in generating the distance slider Omitted atoms can be specified by selecting the appropriate option from the Expert menu and then by clicking on the relevant atoms in the resulting pop up display Conformationally flexible central groups may also be dealt with by means of the Flexible atoms option described next Flexible atoms When a scatterplot is generated the various central group fragments found in the ConQuest search must be superimposed and then averaged Sometimes a central group will contain a conformationally flexible moiety such as a rotationally mobile methyl group Inclusion of these moieties in the superposition process can result in distorted average central group geometries In such cases the user can designate the conformationally variable atoms as flexible IsoGen will keep a separate atomic position for every flexible atom in the ConQuest output file For example here is a plot of carbonyl groups around a mesityl central group The hydrogen atoms of the mesityl methyl groups are designated as flexible Hence
7. e Contour colours can be changed by clicking on the Color button adjacent to each contour This opens a palette from which any desired colour may be chosen e Once all the levels and colours are as you want them click the Create button The contours at the specified levels and colours will appear in the 3D view TsoStar User Guide 21 E Contour Surfaces Create New IsoStar Contour Surfaces C Internal Scaling 0 lt Level lt 100 External Scaling 0 lt Level Create 3 gt new surface s using Probe carbon Color om Level 2 Color _ E Level fi Color _ E Level fos Create m Change Level Color or Visibility of Existing Surfaces for Current Scatterplot Current Scatterplot CSD aromatic iodo any C O 75 000 Internal a Id 50 000 Internal a y 25 000 Internal e Ifyou want to see the surface but not the contacts the latter may be hidden by switching on the Hide All check box which is near the top right of the main graphical interface e The bottom of the Contour Surfaces dialogue lists the contours that have been generated their levels and their colours Change Level Color or Visibility of Existing Surfaces for Current Scatterplot Current Scatterplot CSD aromatic iodo any C 0 2 E men ternal 4 Internal Internal H Delete e Several options are available from within the Contour Surfaces dialogue the display updates automat
8. structures including many protein ligand complexes Only protein ligand ligand water and protein water contacts are included in IsoStar scatterplots 1 e contacts between protein atoms are ignored A subset of the database is used for IsoStar Amongst the restrictions applied are e Crystallographic determinations only no NMR structures e Resolution less than or equal to 2 5A e Protein ligand complexes only A ligand is defined as a peptide of up to ten residues or a non peptide or non haem molecule of at least nine atoms e DNA and RNA complexes excluded e Most covalently bonded complexes excluded e Most duplicate and near duplicate structures excluded e g structures with the words mutant mutation or mutated in their PDB files e No ligands with atoms of partial occupancy e For structures containing more than one binding site with the same ligand only one of the binding sites is used To generate the IsoStar plots ligands of interest and their surrounding residues ie binding sites are identified in all structures Crystallographic symmetry operators are applied to generate the full ligand environment where appropriate e In addition a few structures have been eliminated because they caused processing problems TsoStar User Guide This still leaves several thousand protein ligand complexes for use in building IsoStar 6 1 3 Nonbonded Contact Definition van der Waals Radii e Only intermolecular contacts are inc
9. 4 3 IsoGen Top Level Menu Options e A number of top level menu options are provided e File see Section 4 3 1 e View see Section 4 3 2 e These are only activated when the program has sufficient information for them to be relevant For example the Run button will only be active if a valid cor file 1s named in the Coordinate file box 4 3 1 File Load new search Load settings Save settings Exit bocen The File menu displays a sub menu with the following options e Load new search this command allows the user to load a new set of files from a ConQuest nonbonded search The program will automatically read the appropriate ConQuest output files and generate a new set of defaults e g for central group and contact group atom numbers If a file called lt search name gt rules exists then the program will prompt the user as to whether it should use this file or re generate the settings from fresh see next item for more information on settings e Load settings when a scatterplot is successfully calculated by IsoGen details of the scatterplot settings e g 3D symmetry etc can be saved in a file called lt search name gt rules The Load Settings command allows the user to load a settings i e rules file saved in a previous run of IsoGen e Save settings this command allows the user to save the current scatterplot settings e g containing details of 3D symmetry etc in a settings file which should have the extension
10. After the plot is calculated the user will be given the option of viewing it in the usual way i e using the IsoStar graphical interface e To get help when using IsoGen click with the right hand mouse button on the item for which information is required 4 2 1 Isostar file name e The filename to which the results of the superposition 1 e the scatterplot are written This should have the extension istr the istr format is closely related to the mo 2 format of Tripos Inc Tripos Associates St Louis Missouri USA 4 2 2 Central group atoms e The atoms of the central group 1 e the fragment which will be superimposed The atom numbers are taken from the ConQuest search There must be at least three non linear atoms in the central group 4 2 3 Contact group atoms e The atoms of the contact group 1 e the fragment whose distribution around the central group will be shown The atom numbers are taken from the ConQuest search 4 2 4 Allow short contacts e If this button is switched on all contacts will be included in the scatterplot irrespective of their distance If the button is switched off very short contacts viz more than 1 3 A shorter than the sum of van der Waals radii will be excluded 4 2 5 Switch groups e This command transposes the contact group and the central group It is only enabled when there are sufficient atoms in the contact group for superposition 1 e at least three TsoStar User Guide 59
11. V sum of van der Waals radii of O O or O H This is illustrated in the slider controls at the top right of the IsoStar graphical interface Show contacts in van der Waals vd corrected distance range Lower Limit AOAODODODODODODONONODODONONONODONONONONONONONONONONONONONOONONONONONONONONONONONONONONONONONOUONONONOUONONONOUONOOOOOOOOUOUOOOOOOOOONOOONOUOOOOOOOUOOOOOOOO OOOO ON ONONO N MI 1 21 SSS Sa a Upper Limit POPUP CECE CECE CECE CECE CECE CECE EE EE AD ADDED ADEE AE AD TEE EEE CEE ONDO ONON ANON ANNIN ey 0 5 Hide All Overview vd Over laps Show All e Hit the Overview button The resultant dialogue lists all the CSD reference codes for the contact groups in the plot The distances are reported based on how much shorter the specific contact is compared to vdW radii thus the shortest contact FIDVUK is 1 21 A shorter than the sum of van der Waals radii Once you have finished inspecting the contact groups close the dialogue e Hit the button labelled vAW Overlaps This removes all contacts that are longer than the sum of van der Waals radii from the scatterplot Hitting the button marked Show AII will re display the longer contacts e Alternatively contacts can be removed by using the slider arrows The UpperLimit arrow will remove longer contacts while the LowerLimit arrow will remove short contacts As the slider is moved the two white boxes next to the slider update to reflect the range of contact distances on displ
12. any OH Current visualiser Filename C DOCUME 1 ROBERT 1 LOCALS 1 Temp getting_started_csd_plot_iistr 4 Hyperlink to a CSD Structure e If we want to know more about this unusually short contact we can hyperlink to the original CSD entry to view further details Activate the Hyperlink tick box above the 3D visualiser then click on the either the H or the O atom of the hydroxyl contact group The contact will turn green indicating it has been selected and the hyperlinked plot will be loaded into the second visualiser window In addition the tabbed view in the top left of the graphical interface has changed from Scatterplots to Hyperlinks and the CSD identifier FIDVUK is given in the CSD Identifier window 84 IsoStar User Guide C IsoStar Sm E3 File Options Help Previous Files Show contacts in van der Waals vd corrected distance range Scatterplots Hyperlinks Theoretical Energy Minima Lower Limit ara N E EEE EEE e EE eE 0 6 Identifier Contact number Scatterplot SS aaa esp FIDVUK 101 alhatc alohatc che Info Upper Limit SS eee f 1 21 PDB zi Ena Hide All Overview a vd Overlaps Show All Contour Surfaces Show All Hide All Create Edit Scatterplot Symmetry Expand Contract V Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab Scatterplot CSD aliphatic aliphatic ether any OH Current visualiser Filename C DOCUME ROBER
13. displays all minima e g 4 2 Statistical Data e Some simple statistics are included in IsoStar They can be displayed by navigating to the web browser page for the central group of interest see Section 2 6 and clicking on one of the blue buttons in the Stats column e g TsoStar User Guide 25 General C H only N H O H Other N or O Sulfur Halo halide Amino acid O H Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any OH 2429 207 d alcohol OH 628 22 amp phenol OH 564 21 d water 885 164 al This search has not been done e The following example is taken from a typical statistics page furan any OH Statistics for contacts between furan and any m More Info on the Statistics Source Nb Ne 100 Ne Nb drel CSD 335 22 36 4 0 6 0 1 e The statistics include e The number of crystal structures in the CSD that contain both groups irrespective of whether they form a contact In the above example this would be the number of CSD structures containing both a furan ring and a hydroxyl group e The number of crystal structures N in which the two groups form at least one intermolecular interaction that 1s shorter than the sum of van der Waals radu In the above example this would be the number of CSD structures containing a furan ring and a hydroxyl group in van der Waals contact
14. e The ratio N N expressed as a percentage The bigger the ratio the more often the groups form short interactions Obviously the ratio says something about the extent to which the groups attract each other However it is only a very crude measure as it does not take stoichiometric factors into account For example N N tends to be high when the contact group is C ar H simply because most structures contain many C ar H groupings An estimated standard deviation for the ratio is given in brackets assuming Poisson statistics e The relative density d This is a somewhat less crude measure of the tendency for the groups to form short interactions It is defined as d e dshort dlongs Where dspo 7 18 the density of contacts within the sum of van der Waals radu V and diong 1s the density of contacts between V and V 7ol The larger d 1s the greater the tendency for the groups to form short TsoStar User Guide interactions Typical values for d o are 2 8 for a strong hydrogen bond e g an H bond where one or both of the groups is are charged 1 2 for an average hydrogen bond 0 7 1 for a weak H bond and 0 3 0 7 for a very weak attractive interaction An estimated standard deviation for d e is given in brackets assuming Poisson statistics TsoStar User Guide 27 5 Using the 3D Visualiser 5 1 Controlling which Visualiser is Used e The IsoStar graphical interface contains two visualisers At any given time one of them
15. which are not included in the standard IsoStar library Plots calculated by users with IsoGen can be viewed in the IsoStar graphical interface e Two restrictions should be noted First IsoGen can only be used to calculate scatterplots from the Cambridge Structural Database CSD not from the Protein Data Bank PDB Secondly the external scaling method of contouring density surfaces is unavailable for user generated scatterplots e There are three main stages in producing a scatterplot with IsoGen e A ConQuest search must first be performed to find structures in the CSD that contain the nonbonded contact of interest The crystallographic fractional coordinates of the hits must be saved as a cor file e IsoGen must then be run using the cor tab fgn and fed files from the ConQuest search as input Usually all the default settings of the program can be used all the user has to do is identify the relevant search name and then pick Run from the top level IsoGen menu The program will read the nonbonded fragments from the cor file superimpose the central group detect and eliminate conformational outliers identify and deal with symmetry and calculate the scatterplot e The plot may then be visualised and manipulated in the normal way using the IsoStar graphical interface e A Custom Plots directory is included in the IsoStar directory structure for storage of user generated scatterplots Assuming file protections are set appropr
16. Nn kh Ww NY IsoGen Quick Summary Introduction to IsoGen Nonbonded Searching Using ConQuest Isogen Basic Features Isogen Expert Features IsoGen Theory The Scatterplot Calculation Procedure TsoStar User Guide TIsoStar User Guide 1 Introduction 1 1 Overview of IsoStar e IsoStar is a library of information about the intermolecular interactions formed by a wide variety of chemical groups It is designed primarily for e molecular modellers engaged in structure based drug design e medicinal chemists interested in identifying bioisosteric replacements e protein crystallographers e crystal engineers e IsoStar contains data on e intermolecular interactions in small molecule crystal structures taken from the Cambridge Structural Database CSD e protein ligand interactions in X ray structures from the Protein Data Bank PDB e theoretical intermolecular interaction energies calculated by the Intermolecular Perturbation Theory IMPT method e Crystallographic information in IsoStar is presented as 3D scatterplots Each scatterplot has been calculated by searching the CSD or PDB for nonbonded interactions between a pair of functional groups A and B The A B contacts are transformed so that the A groups are least squared superimposed The resulting scatterplot shows the experimentally observed distribution of B the contact group around A the central group e g e Such a scatterplot gives information about the fre
17. Object 1 Select atoms main window or object list below Current Selection Valid Parameters Defined Objects CENTI s i Valid Objects Centroia Deal RingMaker al 4 eee Delete e Now hit the N and H atoms of the N H fragment and then pick Define next to Centroid again A second centroid CENT 2 is now defined 104 TsoStar User Guide Draw 1 New File Edit Atoms Bonds 3D Options Help ADD 3D CONTACT RingMaker Templates e Next we need to define the actual search _ Select atoms to define 3D Parameters or Objects 10 a Groups ie A Options Delete x To construct Parameter Object Select atoms main window or object list below All Parameters Current Selection Defined Objects Valid Parameters CENT1 CENT2 Valid Objects Centroid O CENT v Delete Reset Done e Select both CENT1 and CENT2 from the Defined Objects box then click on the Define box next to Distance A Distance Type window will appear select the Contact radio button then click OK In the resultant window ensure that the contact is defined between the two centroids and click Define e We will now have to define a contact distance TsoStar User Guide 105 Fie Edit Atoms Bonds 3D Options Help Contact dialog box displayed
18. Objects CENT1 CENT2 Valid Objects Centroid e Leave the Geometric Parameters window open and select both CENT1 and CENT2 in the Defined Objects window Then click on Define next to Distance e A Distance Type window will pop up CHOOSE DISTANCE TPE Intramolecular Distance Select this option to calculate distances between these fragments but only if they are in the same molecule f Contact Select this option to find non bonded interactions between these fragments C Any IF you select this option then the distance calculated map be arbritarily calculated for some entries unless you define a Contact or Intramolecular distance between these fragments Cancel e Select the radio button next to Contact and click OK TsoStar User Guide 57 e In the resultant window click on the Define button This will allow you to input a distance range for the contact e g 0 0 to 3 0 Once this has been done the search can be run 3 2 3 Correcting Hydrogen Atom Positions e Hydrogen X H valence bond distances are usually too short when measured by X ray diffraction To correct for this systematic error ensure the Normalise terminal H positions tickbox is turned on under the Advanced Options tab in the Search Setup window 4 Isogen Basic Features 4 1 Starting IsoGen e Once a ConQuest nonbonded search has been run and assuming IsoStar is installed the IsoGen program can be started by typing
19. Section 5 1 and for reloading a scatterplot that has already been viewed see Section 5 2 3 2 2 Displaying Contacts in a Particular Distance Range e The top right of the IsoStar graphical interface contains options that can be used when the current visualiser see Section 5 1 contains a scatterplot They allow control over which contacts are displayed Show contacts in van der Waals vdi corrected distance range Lower Limit CUPP UPC UECE ACE OE ACE OCE ODEO CE OEE OP COPE OED AE CAE DE EOE ACE EEE A TEASE OEE OE EOE E OEE OE EOE OEE EEE 1 12 r Upper Limit AOAAODOONODOODOOOOODOONOONOONONOONOONOOOOONOONOAOOOOOOOOOOONOON MNOME M EM o s Hide All Overview A vd Overlaps Show All TsoStar User Guide 15 By default the scatterplot will show all contacts shorter than V 0 5 A where V is the sum of the van der Waals radii see Section 6 1 3 of the atoms involved Hitting the vdW overlaps button hides all contact groups except for those within van der Waals contact of any target atom see Section 6 1 4 in the central group in other words you will see only the van der Waals clashes The Overview button lists the CSD reference codes for all the contact groups in the scatterplot ordered from shortest contact to longest Also available are statistics relating to the scatterplot see Section 3 2 3 Hitting Show All will display all contacts Switching on the Hide All check box will hide all contacts this is useful
20. acid trans You should find that all three numbers are the same e The reason is that the three groups carboxylate carboxylic acid cis and carboxylic acid trans cannot be distinguished in protein X ray structures because the positions of hydrogen atoms are not located Hence the PDB data for these three groups are merged together Thus for any given contact group e g water you will get the same scatterplot whether you select from the charged uncharged cis or uncharged trans contact group tables TsoStar User Guide 99 Return to the uncharged carboxylic acid cis page by clicking on the appropriate hyperlink at the top of the Web page 16 Ligand and Protein Central Groups 100 There 1s one more important point to be made about PDB data Look at the menu on the left hand side of the Web page The central groups are divided into two main sections Ligand and Protein Carboxylate can be found in both sections but the PDB based plots in the two sections are not the same This is because the Ligand section contains contacts to carboxylate groups on ligand molecules However the plots in the Protein section show contacts to the carboxylate groups of Asp and Glu sidechains You are currently in the Ligand section of the IsoStar library Click on any number in the PDB column of the contact group table e g amide N H to display a PDB based scatterplot Now hyperlink from any contact atoms in the scatterplot A PDB reference co
21. amide NH as before e Hyperlink from any contact in the new scatterplot A PDB reference code will appear in the PDB Identifier window and the hyperlinked plot will be displayed in the second visualiser window e The resulting display will show a non bonded contact to the carboxylic acid moiety of an Asp or Glu side chain on the protein Remember you are now in the Protein section of the library e This illustrates that for the PDB based data all the scatterplots in the Ligand section of IsoStar show contacts to functional groups on small i e ligand molecules Conversely all the scatterplots in the Protein section show contacts to functional groups on protein side chains or the peptide linkages of protein backbones Strictly speaking all the CSD based scatterplots should be put under the Ligand section of IsoStar since the CSD contains only small molecules not proteins However CSD data have been included in the Protein section as well so that comparisons can easily be made between nonbonded contacts to protein functional groups and contacts to similar groups in small molecules This ends the tutorial TsoStar User Guide 101 Tutorial 3 Calculating Customised Scatterplots Using IsoGen UNIX only Nm hk WO NN 7 The Example Setting up a Nonbonded Search of the CSD Setting up 3D Constraints Running the Nonbonded Search Calculating a Scatterplot Displaying Torsional Flexibility Omitting Atoms 1 The Example e
22. ccdc cam ac uk products life_sciences hermes e mol2 the scatterplot will be saved in mol2 format i e the Tripos Inc format for 3D molecules and crystal structures e pdb the scatterplot will be saved in the Protein Data Bank s format for 3D molecules e jpg a graphical image of the scatterplot as it appears in the 3D display will be saved in jpg format e All atoms will be written out including any that are hidden i e currently not displayed because of the settings that have been chosen for the distance slider TsoStar User Guide 23 4 Viewing Other Data 4 1 Theoretical Data e IsoStar contains the results of theoretical energy calculations performed on various model systems Each system comprises two molecules that model a particular intermolecular interaction For example the interaction between the central group C sp COC sp e aliphatic ketone and the contact group NH amide has been modelled by the molecular complex acetone N methylacetamide Using a model system such as this relatively quick calculations are first performed to search the potential energy hypersurface of the dimer This enables low energy orientations to be located Higher quality ab initio based intermolecular energies are then computed for these low energy dimer orientations using intermolecular perturbation theory see Section 6 3 e Theoretical results are associated with the scatterplot see Section 3 1 to which they are most rel
23. click anywhere in the visualiser and select Measure or Picking Mode followed by Default Picking Mode To clear all measurements from the display right click anywhere in the visualiser and select Clear Measurements TsoStar User Guide 3 6 Experimental Details and Limitations 6 1 Details of Scatterplot Methodology 6 1 1 Structures Used for Calculating CSD Based Scatterplots e The Cambridge Structural Database CSD hAttp www ccdc cam ac uk products csd contains the results of organic and metallo organic crystal structure determinations Standard software ConQuest can be used to search the database for intermolecular contacts between any specified pair of chemical groups If the structure of a compound occurs more than once in the CSD duplicate structure determinations of the same crystal form or determinations of different crystal forms only one example is included in the searches used to set up the IsoStar scatterplots With this exception all CSD structures are used for the IsoStar searches regardless of crystallographic R factor presence of disorder etc However CSD structures with missing hydrogen atoms are not used in creating IsoStar scatterplots unless the missing hydrogen atoms are irrelevant to the plot 1 e not part of either the central or contact group 6 1 2 Structures Used for Calculating PDB Based Scatterplots 32 e The Protein Data Bank PDB http www rcsb org pdb contains the results of protein
24. energy TsoStar User Guide orientation of fluorobenzene N methylacetamide involves an N H F hydrogen bond shown below Such an interaction 1s extremely unlikely to occur in condensed phases where better hydrogen bond acceptors will successfully compete for the NH proton e Calculated interaction energies are always much larger i e more negative for model systems in which one or both of the interacting species is an ion Again this is a consequence of the gas phase nature of the calculations e The value of the MO results is that they can provide independent evidence that the features seen on CSD and PDB scatterplots reflect the intrinsic preferences of the interacting species rather than say crystal packing effects A good example is provided by the phenomenon of H bond lone pair directionality to carboxylate oxygens 7 Example Results 7 1 Carboxylate OH e Many hydrogen bonding groups are included in the IsoStar library Inspection of the CSD based plot of OH around carboxylate shows a distinct preference for H bonding in the oxygen lone pair directions The preference is somewhat obscured when all the contact groups are displayed but it becomes clear if the distance slider 1s manipulated so that only short contacts are shown i e shorter than the sum of van der Waals radii 0 9 A TsoStar User Guide 43 e The preference for lone pair directions becomes obvious if the scatterplot is converted to a contoured den
25. every individual methyl group in the ConQuest file is shown explicitly Flexible atoms are not used in the central group superposition calculation TsoStar User Guide 5 6 5 7 Flexible atoms can be specified by selecting the appropriate option from the Expert menu and then by clicking on the desired atoms in the resulting pop up display Contour probes A contour probe is an atom of the contact group from which it is possible to compute a contoured density surface in IsoStar For example if the contact group were carbonyl it would normally be possible to generate surfaces based on the positions of 1 the carbonyl carbons or 2 the carbonyl oxygens in this situation both the carbon and oxygen atoms are therefore contour probes By default all contact group atoms will be defined as contour probes by IsoGen Each probe atom will be assigned the label used in the ConQuest search The definition and labelling of contour probes can be changed by using the Contour probes option of the Expert menu Selection of this option will delete the default definitions of contour probes and allow the user to select and label individual atoms These atoms alone can then be used for the calculation of contoured density surfaces Use dummy atoms in fitting Sometimes a central group will contain one or more dummy atoms This is usually when a variable element type has been defined in the ConQuest search If the Use dummy atoms in fitting butt
26. hydrogen atoms bonded to rings e g the H atoms in the 4 and 5 positions above are shown 6 1 5 Treatment of Hydrogen Atom Positions e X H covalent bond lengths in IsoStar scatterplots are normalised Normalisation of X H distances involves moving the H atom along the X H vector until the X H distance 1s equal to standard values C H 1 083 A N H 1 009 A O H 0 983 A This corrects for the systematic underestimation of X H bond lengths by X ray diffraction 6 1 6 Constructing Scatterplots when Central Group Hydrogen Atoms are Missing 34 e Insome of the CSD based scatterplots the central group contains only three atoms one of which is a hydrogen e g C sp OH In such cases the hydrogen atom must be used for overlaying when the scatterplot is constructed Clearly this 1s not possible with PDB data because the H atom coordinates will be missing In these cases an extra non hydrogen atom is therefore added to the central group e g the CSD central group C sp OH is changed to C CH OH for the PDB plot The scatterplot below shows the distribution of oxygen atoms around an aliphatic alcohol C sp OH CSD based all contacts are shorter than the sum of van der Waals radii TsoStar User Guide e The scatterplot below shows the distribution of oxygen atoms around an aliphatic alcohol C CH O H PDB based all contacts are shorter than the sum of van der Waals radii e The absence of H atoms in protein str
27. if you want to create a picture of a central group with a contoured density surface see Section 3 4 The sliders allow interactive control over which contacts are displayed If the top Lower Limit slider is moved to the right short contacts will be hidden from the display If the bottom Upper Limit slider 1s moved to the left long contacts will be hidden At any given slider setting the top and bottom white boxes show the limits of the current display For example with the sliders in the positions shown below all contacts in the distance range V 0 1 to V 0 2 A are displayed where again V is the sum of the van der Waals radii of the atoms involved Show contacts in van der Waals vd j corrected distance range Lower Limit F Upper Limit T Hide All Overview a wily a Show All m You can type distance limits directly into the white boxes but you must terminate each number with a carriage return 1 e Enter for it to take effect Moving the middle grey rectangle with the mouse alters both sliders simultaneously By setting a narrow range e g the above setting corresponds to a range of 0 3A this therefore allows you to slice through the contacts 3 2 3 Viewing Scatterplot Contact Distributions 16 e Contact distribution histogram plots of the scatterplot as well as a list of refcodes that make up the plot ordered by shortest to longest contact distance are provided under the Overview button TsoSta
28. requested e Reduction of distribution into an asymmetric unit e Writing of output files Further details of some of the methods used are summarised in the sections that follow e Derivation of Topological Symmetry see Section 6 2 e Derivation of Initial Model see Section 6 3 e Superposition see Section 6 4 e Derivation of 3D Symmetry see Section 6 5 Derivation of Topological Symmetry Some chemical groups contain atoms that are topologically equivalent whilst not being symmetry equivalent in three dimensions An example of such a fragment would be vinyl TIsoStar User Guide H2 e Here the two hydrogen atoms H and H2 are not symmetry related in 3D but are topologically equivalent in 2D 1 e the CH group has localised mm2 symmetry As a result the matching of HI and H2 onto the atoms of the ConQuest search query is arbitrary The consequence of this is that the Z hydrogen H2 and the E hydrogen H7 will appear in an arbitrary order in the ConQuest output files IsoGen must detect this in order to achieve correct superposition of central group fragments since clearly it must not attempt to fit the Z hydrogen of one vinyl fragment onto the E hydrogen of another e Topologically equivalent atoms are detected by using a modified version of the Morgan algorithm Morgan 1965 An explanation of this algorithm is provided later see Appendix 2 Algorithm for Identifying Topological Equivalence page 77 6 3 Derivation
29. see Section 4 1 IsoStar User Guide ll 3 Viewing and Using Scatterplots 3 1 Introduction to Scatterplots in IsoStar e The presentation of nonbonded contacts as 3D scatterplots is best explained by an example Suppose we are interested in the interaction of aromatic iodo substituents C ar I with oxygen atoms Using standard software ConQuest the CSD can be searched to find crystal structures containing C ar I groups that form 1odine oxygen nonbonded contacts shorter than some specified distance we use sum of van der Waals radii 0 5A Two of the first hits found in this search are I 0 contacts in CSD entries ACICOD and AJBFOR10 e The 1odine oxygen fragments are extracted from each structure including the first three carbon atoms of the aromatic ring The iodine and ring atoms of one fragment are then overlaid on those from the other to give a composite picture in which the I O contacts from the two structures are displayed simultaneously relative to the same frame of reference 12 TsoStar User Guide e Repetition of this process for all hits found in the CSD produces a scatterplot showing how oxygens are distributed around aromatic iodo substituents in small molecule crystal structures e In IsoStar this scatterplot is displayed in three dimensions and is symmetrised i e all the contacts are reflected into one quadrant since the aromatic iodine group has two planes of mirror symmetry A scatterplot showing
30. the distribution of OH ether contacts as observed in protein ligand complexes from the PDB Because hydrogen atoms are not located in protein crystal structures TsoStar User Guide 87 this plot does not show the H atoms of the hydroxyl groups just the O atoms e By comparing the PDB plot with the CSD plot you can see that both distributions are similar geometrically 7 Hyperlink to a PDB Structure e Move the Upper Limit slider to the far left to display the shortest contact e Activate hyperlinking by clicking on the Hyperlink Mode tick box then select the remaining single contact atom in the 3D view The contact will turn green and the hyperlinked structure will appear in the second visualiser window SS TsoStar User Guide Hyperlink FDB 103401 pam A y kyi REGIO l f WA O Current visualiser lt Back Fuyv gt e The PDB code of the structure containing this short contact 1QA3 is displayed in the PDB idenfier window Details of the structure including publication details are provided under the Info button EE PDB Hyperlink Information PDB Code 1034 Molecule STROMELYSIN 2 CALDERONE IBERTINI M FRAGAI C LUCHINAT S MANGANI B TERNI Author s J MOL BIOL 336 707 2004 Reference e Inspect the active site in the hyperlinked structure It shows the ligand together with nearby water molecules and protein residues In fact the contact 1s between the active site ligand and a TsoSt
31. ules e Exit IsoGen this command exits IsoGen 4 3 2 View Search fragment Current Bo star file Ex pert options 60 TsoStar User Guide The View menu displays a sub menu with the following options e Search fragment this command allows the user to view the search query as drawn in ConQuest and with ConQuest atom numbers displayed This option will be greyed out if there are no 2D coordinates in the ConQuest output files e Current IsoStar file this command allows the user to view the current scatterplot It is greyed out if no istr file exists e Expert options this command gives the user access to a number of expert options see Section 5 that are available in IsoGen Most of these options can also be accessed through the top level menu item Expert e Expert gives access to various expert options see Section 5 e Run initiates calculation of the scatterplot In most cases the IsoGen program defaults are adequate to produce an acceptable scatterplot In such cases clicking on Run in the top level menu is all the user need do once the IsoGen interface has been opened After the plot has been calculated the user will be given the option of viewing it in the usual way i e using the IsoStar graphical interface e Help displays a sub menu with the following options e Getting help displays a pop up window which explains the ways of getting help in IsoGen 1 e via the main Help document or via context dependent he
32. 828 A In this way we get corrected distances of e O4 H5 0 828 A 04 06 0 331 A C2 H5 0 004 A C2 06 0 381 A e Now suppose that C2 04 H6 and O6 were all slider atoms and the distance slider range in IsoStar was set to display all contacts between 1 0 and 0 5 In this case the contact described above would be displayed because the shortest corrected distance between a central group slider atom and a contact group slider atom 1 e 0 828 falls in the range 1 0 to 0 5 However if only C2 O4 and O6 were slider atoms the contact would not be displayed because the shortest distance between slider atoms 0 331 would now fall outside the specified range e By default any atom utilised in a ConQuest CONTACT search is designated as a slider atom The default settings can be changed by means of the Slider atoms option This opens a pop up window containing a display of the ConQuest search query the user may then click on the desired slider atoms 5 4 Omit atoms e When a scatterplot is generated the various central group fragments found in the ConQuest search must be superimposed and then averaged Sometimes a central group will contain a conformationally flexible moiety such as a rotationally mobile methyl group Inclusion of these moieties in the superposition process can result in distorted average central group geometries e In such cases the user may wish to omit the atoms whose positions are conformationally
33. Although there is an extensive range of scatterplots in IsoStar there will probably be occasions when you want to look at a plot that is not in the library This tutorial will demonstrate how to perform a nonbonded search of the CSD using ConQuest and then convert the results into a scatterplot We will take as an example the methylcarbamoyl group CONHMe and look at how it forms hydrogen bonds It is assumed that you have basic familiarity with ConQuest specifically how to build fragments and run simple substructure searches If not you should read the ConQuest documentation or use the ConQuest tutorials available under the Help menu or at the end of the documentation 2 Setting up a Nonbonded Search of the CSD e Open ConQuest and launch the sketcher window by clicking on Draw in the main interface e Draw the methylcarbamoyl fragment C CONHMe Include the H atoms and the carbon atom to which the methylcarbamoyl fragment is attached 102 TIsoStar User Guide Draw 1 New heal x File Edit Atoms Bonds 3D Options Help oo Click and PREE ee eat rea to create a bond Next Atom H 2 3D Parameters as al PREE ee eat rea to an existing atom to make a connection Next Bond Single KAA Options CONTACT Delete Contacts Options Delete RingMaker Search biw CTH o N s P F CI An More Grou z a Bond ngl Ei Templates E E Store Cancel e Now draw a separate N H group Dra
34. CT Dale oO C C Q RingMaker Search 4 rir 4 Store CA BR O N S P cl Any Mare Groups C Bond Single md Templates Cancel e The next step is to define a contact between the oxygen atom of the ketone group and the hydrogen of the O H group using the CONTACT button in the sketcher TsoStar User Guide 55 O x File Edit Atoms Bonds 3D Options Help Select atoms to define non bonded contact or group Defined Contact CON1 Inter Ydw 0 04 F 3D Parame ters Atom 04 to Atom HE DRAW EDIT ERASE ADD 3D Options CONTACT Delete Contacts 5 CON1 04HE Pa 64 Options Delete Tn ed Contact Defir d 1 C C CONTACT CONI Hit Edit to change settings or select atoms to create additional contact VU ATOM 04 ATOM H6 Defined Contact 04 H6 e a Ced Inter molecular Ring al Z yy ell S acme ll Distance within sum of Ydw 0 04 HAQM AKELr e Once the contact has been defined the search can be started by clicking on Search e It 1s important that the Normalise terminal H positions tickbox is turned on under the Advanced Options tab in the Search Setup window e The search can be started by clicking Start Search 3 2 Advanced Nonbonded Search Options 3 2 1 Defining Distance Ranges e When a contact is defined in ConQuest the default distance of less than the sum of Van der Waals radii are used un
35. IsoStar User Guide and Tutorials 2010 Release Copyright 2009 The Cambridge Crystallographic Data Centre Registered Charity No 800579 Conditions of Use The Cambridge Structural Database System CSD System comprising all or some of the following ConQuest Quest PreQuest Mercury Mercury CSD and Materials module of Mercury VISTA Mogul IsoStar SuperStar web accessible CSD tools and services WebCSD CSD Java sketcher CSD data file CSD UNITY CSD MDL CSD SDfile CSD data updates sub files derived from the foregoing data files documentation and command procedures each individually a Component is a database and copyright work belonging to the Cambridge Crystallographic Data Centre CCDC and its licensors and all rights are protected Use of the CSD System is permitted solely in accordance with a valid Licence of Access Agreement and all Components included are proprietary When a Component is supplied independently of the CSD System its use 1s subject to the conditions of the separate licence All persons accessing the CSD System or its Components should make themselves aware of the conditions contained in the Licence of Access Agreement or the relevant licence In particular e The CSD System and its Components are licensed subject to a time limit for use by a specified organisation at a specified location e The CSD System and its Components are to be treated as confidential and may NOT be disclosed or re distributed in any fo
36. Star User Guide
37. T ILOCALS 1 Temp getting_started_csd_plot_Listr e Ensure the hyperlinked display is set as the Current visualiser either by clicking anywhere in the 3D view clicking on the greyed out Hyperlink CSD FIDVUK button at the top of the hyperlinked display or by hitting the Current visualiser radio button at the bottom of the hyperlinked display e The fragments involved in the contact are highlighted in ball and stick mode e Measure the contact distance by right clicking in the 3D display selecting Measure and Measure Distances from the pull down menu then clicking on the ether O and hydroxyl H atoms The distance will be reported as 1 563 A e Hit the Info button to the right of the CSD Identifier display This launches a dialogue that provides further information about the structure e g the compound name literature reference and crystallographic information Close this dialogue and return to the IsoStar visualiser window e Return to the original scatterplot and display all contacts by clicking on the Show All button and return to the Scatterplots tabbed display TsoStar User Guide 85 5 Generation of Contoured Density Surfaces e Click on the Create Edit button in the Scatterplots tabbed display This launches a Contour 86 Surfaces dialogue which allows you to apply contours to the current scatterplot E Contour Surfaces Fi E Create New IsoStar Contour Surfaces Internal Scaling 0 lt Level lt 100 C External Sc
38. TsoStar User Guide gt IsoStar 2 1 Windows Internet Explorer tn A J Y http intranet ccde cam ac uk isostar Asostar_2 1 htmlfsostar Atml File Edit View Favorites Tools Help Yr 2 v Search 2 A gt PBant spy mal E Answers Dating GJ Mobile I Sign in lsoStar 2 1 Ligand Terminal Ligand Acyclic Links Ligand Ring Systems Ligand Solvates Protein Plots Custom Plots Home Ligand Terminal C H only N C H only O C H only N O C H only Si containing P containing S containing Halo containing Acyclic links C H only r N C H only 4 O C H only i N O C H only P containing S containing Version 2 1 2009 Release Ring systems Phenyls C H only N C H only O C H only N O C H only Welcome to soStar S containing Nucleic acid bases Solvates etc Inorganic Organic sj f T 1 ied Local intranet 100 5 e Although IsoStar contains data for many different functional groups users may want to generate their own scatterplots for contacts not currently included in the library Consequently there is a directory reserved in IsoStar for Custom Plots i e scatterplots created by users with the program IsoGen see IsoGen Quick Summary page 53 e Clicking on one of the central group categories produces a list of available groups e g TsoStar User Guide 9 Ligand Terminal N O C H only e acetylamino e carbamoyl formamido e formamido E
39. act that their positions are usually determined with lower experimental precision 4 Ifany of the original central group atoms cannot be matched with a symmetry generated atom the symmetry element is rejected 5 Ifevery operator for a symmetry element passes the operator is accepted e At the end of the process a symmetry expansion routine is used This computes the products of elements found in the above steps and then tests for further symmetry TsoStar User Guide 75 Appendix 1 Vector Direction Syntax e Vector directions can be specified in IsoGen using the numbers of the atoms from the ConQuest 76 search query So if a ketone group had atoms C1 C2 C3 and O4 with C2 double bonded to O4 and single bonds C1 C2 C3 C2 then e the C2 04 vector would be denoted as 2 4 e the O4 C 2 vector would be denoted as 4 2 e the C1 C3 vector would be denoted as 1 3 etc Vectors can also be referenced with respect to the centroid of the molecule For example the ketone centroid O4 vector would be denoted as c 4 Three operations can be used to combine vectors summation subtraction and cross multiplication For example e The vector sum of the C2 O4 and C2 C3 directions would be denoted 2 4 2 3 each vector is delimited from the operator with a space e A vector perpendicular to the C2 O4 and C2 C3 directions i e their cross product would be denoted 2 4 x 2 3 Currently operator precedence is from left to righ
40. aling 0 lt Level Create 3 EE new surface s using Probe oxygen Color Lom Level 75 Color _ E Level 50 Color E Level 25 Create Change Level Color or Visibility of Existing Surfaces for Current Scatterplot Current Scatterplot CSD aliphatic aliphatic ether any OH Delete e Contour surfaces can be scaled in one of two ways either Internal or External The Internal Scaling radio button will already be activated With Internal scaling the densest region in the scatterplot is assigned a density of 100 and all other regions are scaled relative to this e The number of surfaces and the probe atom can be chosen using the Create new surfaces using Probe pulldowns Note that a maximum of 3 surfaces can be generated Keep thenumber of surfaces at 3 and select hydrogen from the probe pulldown menu e Colours and levels are suggested for the different contours Keep the levels as they are 1 e at 75 50 and 25 The colours can be modified if desired Once the settings are as you want them click Create All three contours wil be added to the Current Scatterplot section of the Contour Surfaces window Change Level Color or Visibility of Existing Surfaces for Current Scatterplot Current Scatterplot CSD aliphatic aliphatic ether any OH 1 hydrogen 75 000 ASE od Internal av hydrogen 25 000 Internal ae lV Delete TsoStar User Guide e From within thi
41. all C ar I O C contacts shorter than sum of van der Waals radii 0 2A is shown below Note that the carbonyl carbon atoms are shown as well as the oxygen atoms that are actually forming the short contacts e It sometimes happens that a particular type of contact occurs so many times in the CSD that the resulting scatterplot 1s very crowded and hard to inspect In such cases a pseudo random subset of contacts is taken to limit the total number of atoms in the scatterplot to about 5 000 e PDB based scatterplots are produced in an analogous way capturing information about protein TsoStar User Guide 13 ligand ligand water and protein water interactions 3 2 Viewing Scatterplots 3 2 1 Loading and Reloading Scatterplots e Use the IsoStar web browser interface to navigate to the table relating to the central group of interest see Section 2 6 e To view a Scatterplot based on CSD information click on a number in the CSD column of the table For example clicking on the number 2486 below will display the scatterplot of nitro H X contacts X O N or S as observed in small molecule crystal structures in the CSD 2486 is the number of contacts in the scatterplot nitro General C H only N H O H Other N or O Sulfur Halo halide Amino acid General 2 Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any C N O S or H 4930 1551 poo any polar X H X N O o
42. almost never located in protein crystal structure analysis causes difficulty in calculating PDB based scatterplots for groups that 1 can exist in different ionisation states 11 have distinct conformations differing only in the positions of one or more hydrogen atoms or 111 have alternative tautomeric forms e The ionisation states of acidic groups and basic groups cannot be determined unambiguously TsoStar User Guide from protein crystal structures They can sometimes be inferred by chemical reasoning but not always and often not with confidence In contrast ionisation states are usually determined reliably in CSD structures Accordingly there are separate CSD based scatterplots in IsoStar for e g carboxylate CO and carboxylic acid CO gt H but only a single combined PDB based plot Since the pK of carboxylic acids is relatively low most of the contributors to the PDB based CO CO gt H plot will be ionised but probably not all e Similarly alternative geometries that differ only in H atom positions cannot be distinguished in protein structures Thus for example there are separate CSD based scatterplots for planar and pyramidal aromatic amino groups but only a single PDB based plot e Tautomeric states are often uncertain in protein structures whereas they are usually determined with confidence in CSD structures For example imidazol 4 yl left below and imidazol 5 yl right below can be distinguished in the CSD bu
43. ar User Guide 5 amp 9 8 water molecule e The contact distance can be measured in the same way as with the CSD plot 1 e by right clicking selecting Measure Distances from the pull down menu and selecting both atoms involved in the contact Inspection of a Theoretical Plot e Now click here to display the minimum energy geometry of the complex between methanol and dimethyl ether e As the two visualiser windows contain the scatterplot and the hyperlinked plot you will obtain an Overwrite Content Warning window asking whether you would like to load the new plot into the Current visualiser 1 e the one with the active Current visualiser radio button or the other visualiser with greyed out headers and footers We are finished with both PDB plots so the new plot can be loaded into either window Click on either Load in Current or Load in Other e This plot was calculated theoretically The number in white is the computed interaction energy between the two molecules in kilojoules mole but note this is a gas phase result e Close the IsoStar client window This finishes the tutorial which has demonstrated most of the IsoStar software Now you might like to work through Tutorial 2 which demonstrates how to find your way through the IsoStar Web pages 90 TIsoStar User Guide Tutorial 2 Navigating the IsoStar Web Pages 8 The Example 9 Selecting a Central Group 10 Displaying CSD Scatterplots 11 Displaying PDB Scatterp
44. ay e For example move the LowerLimit slider arrow until the top box reads 0 60 Now move the UpperLimit slider arrow until the number in the bottom box reads 0 50 All contacts now displayed are within the distance range V 0 6 A to V 0 5 A TsoStar User Guide 83 e Moving the central part of the slider bar enables you to move this 0 1 A display range upwards 1 e up to longer contact distances or downwards towards shorter distances e Specific values may also be typed into the white boxes e Hit the button labelled Show All to re display all contacts Then move the UpperLimit slider arrow as far left as it will go until only one contact the shortest is displayed E Isostar Joj x File Options Help Previous Files Show contacts in van der Waals vd corrected distance range Scatterplots Hyperlinks Theoretical Energy Minima Lower Limit COUPE CECE CUE CE CE CD UDO CE CDE OE CECE DOE EE CEE AE EE CED EE EOE EEOC EEE CEE AE ECE EAE EE EEE EERE EAT OEE ADEE CE CAT AE EEE AD AE PEE AEA EE CE ADOT AEE EAD EERE ONON ON EEE 0 6 CSD Jaliphatic aliphatic ether any OH E ANTUAN TIATIA TTT TTT TTT TITT TITT Upper Limit f 1 21 PDB T Hide All Overview vd Overlaps Show All Contour Surfaces Show All Hide All Create Edit Scatterplot Symmetry Expand Contract IT Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab Scatterplot CSD aliphatic aliphatic ether
45. bonyl groups around ethynyl There is a surprising cluster of oxygens near the tetrahedral carbon e This is due to the fact that many of the CSD structures contain an OH on this carbon 1 e C OH CCH for example CSD entry APYNAC 38 TsoStar User Guide e Secondly CSD based scatterplots may be misleading because of the existence of a common crystal packing motif This is well illustrated by ring systems that contain a cis amide function e g lactams and hydantoins These often dimerise by hydrogen bonding e g e This results in strong clustering on the corresponding scatterplots Thus the distribution of the contact group any C N O S or H around gamma lactams shows clusters of oxygen and nitrogen atoms near the N H and C O groups respectively of the lactam Many though by no means all of these contacts arise from H bonded dimers This is shown in the image below all contacts are shorter than the sum of van der Waals radii 0 1 A TsoStar User Guide 39 6 2 4 Biases in PDB Based Scatterplots e An important source of bias is the number of closely related entries in the PDB For example the distribution of amide carbonyl groups around arginine guanidiniums shows a tight cluster of contacts between the two NH groups all contacts are shorter than the sum of van der Waals radii 0 1 A 4 TsoStar User Guide e On hyperlinking however it is found that most of these contacts come from a series of very closely
46. conformer e formamido Z conformer hydroxyimino e methylcarbamoyl e nitro e Clicking on one of the central group names causes a table to be displayed which is the access point for all data pertaining to that group The screenshot below shows part of this table for nitro 10 TsoStar User Guide nitro General C H only N H O H Other N or O Sulfur Halo halide Amino acid General l Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any C N O S or H 4930 1551 any polar X H X N O or S 2465 386 amp This search has not been done General C H only N H O H Other N or O Sulfur Halo halide Amino acid C H only e Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any alkyl C H 2472 401 methylene 977 129 methyl 999 114 b any aromatic C H 2498 293 substituted aromatic carbon 1194 24 phenyl 686 60 This search has not been done General C H only N H O H Other N or O Sulfur Halo halide Amino acid N H e Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any NH 2390 169 any uncharged NH 2474 s e Clicking on links in this table allows you to e view scatterplots based on CSD or PDB data see Section 3 2 1 e view statistical information on the frequencies of contacts see Section 4 2 e view theoretical results
47. d click OK in the IsoGen text window Most users can finish the tutorial at this point since the key features of ConQuest and IsoGen have now been demonstrated To exit the tutorial close the IsoGen window with File Exit IsoGen However 1f you want to see some Expert IsoGen options proceed to the next page of the TsoStar User Guide tutorial 6 Displaying Torsional Flexibility e We have produced a reasonably good scatterplot using the IsoGen defaults However the methyl group of the methylcarbamoyl fragment adopts a range of torsional conformations and these have been averaged in the scatterplot Suppose we want to show explicitly the existence of the methyl group rotational flexibility We can do this using IsoGen Expert options e Select Expert from the IsoGen top level menu and then pick Flexible atoms from the resulting sub menu A new window will appear which shows the N H and methylcarbamoyl fragments as drawn in ConQuest Pick the three methyl group hydrogen atoms to define them as flexible The window should look something like this but your atom numbers will probably be different as they depend on the order in which you drew the atoms and bonds in ConQuest o Flexible Atoms fog pt F OK Cancel e Hit Ok e Now hit the IsoGen top level menu button Run again A new scatterplot will be calculated Don t worry if you get some warning messages from the program 1f necessary just hit Ok
48. de will appear in the PDB Indentifier window and the hyperlinked structure will appear in the second visualiser window The hyperlinked display shows a protein binding site there 1s a contact between the active site ligand which contains a cis carboxylic acid and a residue in the protein amide containing Close the IsoStar visualiser window graphical interface and return to the browser Scroll down the window until you can see the Protein section Click on Terminal You will obtain a list of terminal chemical groups that occur in the natural amino acids Protein Terminal i e ammonio K Ligand Terminal CHa h Hs CH only NCH only a CCH only MO CH only Si containing P cantaining CH S Containing h Halo containing e carbamoyl N Q Aevelic inks carboxylic acid O E CH only i D MCH only HC Tee only MOCH only s P containing S Containing carboxylic acid DE charged carboxylic acid DE uncharged cis Bas 7 E carboxylic acid uncharged trans e guanidinia H Ring systems Phenyls CH only MH only OCH only NO CH only S Containing Mucleic acid hases hydroxy falipnatic 5 7 Solvetes fc Inorganic TsoStar User Guide e Select any one of the carboxylic acid entries e g carboxylic acid D E uncharged cis from this list The corresponding contact group table will appear e Click on any of the numbers in the PDB column to display a scatterplot e g
49. ed CSD A scatterplot will appear showing the distribution of NH OH and SH groups around carboxylates in the CSD Note that the text at the top of the visualiser window updates to reflect the contents of the visualiser in this case Scatterplot CSD charged carboxylic acid any polar X H X N O or S Again the plot shows that H bonds tend to occur along the oxygen lone pair directions This becomes clear if you move the Lower Limit bar of the slider to the left 1 e display shorter contacts TsoStar User Guide 95 11 Displaying PDB Scatterplots Now go to the contact group table and click on the number in the row marked any polar X H and the column headed PDB In the Overwrite Content Warning window click on the Load in Other button to load the scatterplot into the second visualiser window You will now have the CSD based plot adjacent to the PDB plot The PDB based scatterplot displays the distribution of NH OH and SH groups around carboxylates in PDB protein ligand complexes Since hydrogen atoms are not located in protein crystal structures the plot shows only the non hydrogen atoms Close the IsoStar graphical interface and return to the browser window Look at the contact group table Some of the cell are empty 1 e contain no number This means that no hits were found when the search for that contact was performed If a cell contains a star it means that the relevant search has not yet been done 12 Displaying Statistical Data
50. edit the atom type TsoStar User Guide 67 68 For contact group atoms the atom type is also allowed in IsoGen This makes the program guess the atom type for each individual contact atom an atom type is assigned which gives the atom the correct element symbol although not necessarily the correct hybridisation state Slider atoms A distance slider is provided in the IsoStar graphical interface This allows users to remove from a scatterplot display all contact groups outside a specified corrected distance range the corrected distance between two atoms 1s their interatomic distance minus the sum of their van der Waals radii Only central and contact group atoms defined as s ider atoms are taken into account when IsoStar determines whether a given contact group should be displayed or not given a particular setting of the distance slider The operation of the slider given a particular choice of slider atoms 1s best illustrated by an example Consider the following ConQuest query TsoStar User Guide e Suppose that the first contact found in the ConQuest search had the following interatomic distances e 04 H5 1 892 A 04 06 2 709 A C2 H5 2 896 A C2 06 3 601 A e Each of these values may be converted to a corrected distance by subtracting the relevant van der Waals radii For example the vdw radii of O and H are 1 52 and 1 20 A respectively so the corrected 04 H5 distance 1 892 1 52 1 20 0
51. er or lower than would be expected by chance For example a figure of 2 0 would imply that contacts at that grid point were twice as frequent as would be expected a figure of 0 5 would suggest that the density of contacts was only half the random expectation Contouring 1s then done on these values e The practical difference between internal and external scaling is seen when two different scatterplots are contoured and compared If internal scaling is used the scaling of each 1s purely local so the contour levels of one bear no relation to those of the other If external scaling is used both plots are nominally on the same scale in each case a contour level of x means that contacts are x times as frequent as would be expected by chance 3 5 Saving Scatterplots e To save a scatterplot first ensure that it is being displayed in the current visualiser see Section 5 1 Then select File from the top level menu followed by Save As then select one of the following options from the resultant menu e IsoStar istr this will save the scatterplot in istr format This is basically the Tripos mol2 fomat Tripos Associates St Louis Missouri USA with some extra information held in comment lines Hence it should be possible to read files of this type into a variety of visualisers including the CCDC programs Mercury http www ccdc cam ac uk products csd_system mercury and Hermes the dedicated visualiser provided with Relibase http www
52. erent In this case the two degenerate axes can be placed anywhere in a circular plane Finally in the case where all three radii are equal a sphere the axes can be placed anywhere In mathematical terms this can be regarded as a problem of degeneracy the eigenvalues of the inertia tensor are degenerate The same situation occurs with molecules that have high order symmetry elements To resolve such situations vectors from the molecular centroid to each atom in turn are calculated and tested for symmetry The centroid atom vectors to topologically equivalent atoms are then summed to produce further directions for symmetry testing Additional directions for symmetry testing are calculated by cross multiplying pairs of centroid atom vectors The following algorithm is used to test for a rotational symmetry element of order n around any particular direction Matrix operations are generated for each operator in the symmetry element Each operator is applied in turn to each coordinate in turn A search is made to determine whether the atomic positions generated in step 2 G are equal within defined tolerances to any of the original atomic positions O The tolerances for non hydrogen atoms are 74 e the angle G centroid O is lt 5 0 degrees and e the difference between the G centroid distance and the O centroid distance is lt 0 1 A TsoStar User Guide e Hydrogen atoms have larger tolerances to allow for the f
53. ethanol However the MO calculations predict the opposite lone pair preference to that observed in crystal structures This 1s probably because steric effects are usually more important in the CSD structures than in the very unhindered model system used for the calculations The three lowest potential energy minima of the methyl acetate methanol complex are shown 46 TsoStar User Guide below e Other oxygens in conjugating environments are also poor acceptors For example the CSD based plot of N O S H groups around isoxazole shows that hydrogen bonding almost always occurs to the ring nitrogen not the ring oxygen contacts shorter than the sum of van der Waals radii 0 25 A are shown TsoStar User Guide 47 7 3 Phenyl C O phenyl CH aliphatic e One of the most striking features in IsoStar is the surprisingly strong directional preferences of contacts to hydrophobic groups For example the contoured distribution of carbonyl oxygens around phenyl shows an overwhelming preference for these atoms to occur around the edge of the phenyl ring e In contrast the distribution of aliphatic C H around phenyl shows a peak above the ring e This is consistent with the high quadrupole moment of benzene 48 TsoStar User Guide 7 44 Amidino Carboxylate e In the PDB based plot of carboxylate groups around amidine there are some short contacts above the amidine carbon contacts shorter than the sum of van der Waals radi
54. evant Navigate to the web browser page for the central group of interest see Section 2 6 and click on one of the gold buttons which are only displayed when relevant theoretical results are available e g General C H only N H O H Other N or O Sulfur Halo halide Amino acid N H Links to statistical data D Links to theoretical energy data Contact Group CSD PDB Stats Theory any NH 2390 169 any uncharged NH 2474 h D any cationic NH 1596 14 aromatic cationic N H 343 amide NH 358 52 2 uncharged C sp2 C ar NH2 618 d cationic RNH3 284 14 cationic R3NH 107 This search has not been done e This produces a table listing calculated dimer energies units kJ mol These include the total interaction energy of each geometrical minimum together with a breakdown into the component physical contributions see Section 6 3 e g 24 TsoStar User Guide Theoretical energy data for contacts between aliphatic aliphatic ketone and amide N H Model Compounds Methodology Problems acetone N methyl acetamide interaction Energies kJ mol Minima Fes _ Fer Fpot Fef Sais Etot 1 29 9 22 6 3 4 3 4 10 1 24 1 2 16 8 11 2 1 5 1 0 9 3 17 5 e Calculated minimum energy dimer geometries can be visualised by clicking on one of the numbers in the Minima column displays an individual minimum or clicking on A
55. group are shown since it has one plane of mirror symmetry Scatterplot Symmetry Expand Contract e The symmetry can be expanded if required using the Expand button next to Scatterplot Symmetry e To contract the symmetry 1 e to return the scatterplot to its original settings hit the Contract TsoStar User Guide 17 button 3 2 5 Display of Hydrogen Atoms in Scatterplots e Hydrogen atoms are always included in CSD based scatterplots but never in PDB based plots e This is because hydrogen atoms are almost never located in protein crystal structures whereas usually they are in small molecule structures and when IsoStar scatterplots are created CSD structures will only be used if all the relevant hydrogen atoms positions are available e The lack of hydrogen atoms in PDB based plots sometimes the plots difficult to interpret see Section 6 2 2 3 2 6 Display of Scatterplots for Symmetrical Central Groups e Many of the central groups in IsoStar are symmetrical For these only the asymmetric unit of the scatterplot is shown by default For example the scatterplot of N H around nitro has all the N H groups reflected into one quadrant since the nitro group has mm symmetry e Linear groups such as cyano and ethynyl pose a problem as they have cylindrical symmetry The problem is overcome by including enough extra atoms to break this infinite symmetry For example IsoStar contains C ar CN and C sp CN entries having
56. he aliphatic ketone fragment by default has mm2 symmetry The two fold axis lies along the C O bond A mirror lies in the plane formed by the four atoms This can also be regarded as a two fold rotation inversion axis along any vector perpendicular to the plane Since symmetry definition in IsoGen must be done with respect to a direction not a plane we must regard mirror planes in this way The second mirror plane contains the C O bond and is perpendicular to the first e Only the first two symmetry elements are needed to describe the full point group since the third is generated by the program from the first two The syntax for vector specification is provided later see Appendix 1 Vector Direction Syntax page 76 e The direction of the two fold axis is along the C O bond and so is easily specified 2 JbA e This defines a two fold axis along the bond between atoms 2 and 4 the C O bond in this case e The first mirror plane is specified using the cross product of the two C C bonds the bonds between atoms 2 and and 2 and 3 2 lea m 2 12 e Since the third symmetry element is automatically generated from these two it does not need to be specified explicitly Thus the above input is adequate to describe the mm2 point group 64 IsoStar User Guide 5 1 2 Defining Symmetry Example 2 Benzene e The symmetry of benzene can be described in the following terms e A mirror plane passing through all atoms mirror 1 e A si
57. he scatterplot Reloading Previously Viewed Scatterplots Structures and Minima Scatterplots theoretical energy minima and crystal structures that have been previously loaded in a session may be reloaded in two ways e By use of the Back and Fwd buttons underneath the visualiser these move backwards or forwards through the items that have been loaded into the visualiser Current visualiser lt Back Fwd e By use of the Previous Files options in the top left of the IsoStar graphical interface For example to view a previously loaded PDB based scatterplot hit the Scatterplots tab this may not be necessary if the Scatterplots pane is already at the front and then select the desired PDB scatterplot from the pull down menu FPrevious Files Scatterplots Hyperlinks Theoretical Energy Minima CSD phosphonate any C N O S or H atom PDB phosphonate any alkyl C H x phosphonate any polar lt H NLO or 5 phosphonate any alkyl C H e Access to previously loaded CSD scatterplots can be obtained via the CSD pull down menu and the Hyperlinks and Theoretical Energy Minima tabs similarly give access to pull down menus of previously loaded crystal structures and energy minima Rotating Translating and Scaling Rotate by moving the cursor around in the visualiser while keeping the left hand mouse button pressed down Rotate around the z axis the axis perpendicular to the
58. his file with the same name as before e g CONHMe cor e The new directory should now contain 4 files e CONHMe tab e CONHMe fgn e CONHMe fed TsoStar User Guide 107 108 e CONHMe cor Browse to the directory containing all the files and open Isogen by typing the following at the command prompt 1sogen CONHMe amp You should now have a control window for a program called IsoGen which reads the cor and jnl files and constructs an IsoStar like scatterplot In most cases including this one it is possible to calculate a reasonable scatterplot using the standard IsoGen defaults The program will automatically detect topologically symmetrical atoms overall fragment symmetry conformational outliers etc So just hit the IsoGen top level menu option Run A new window will appear which will keep you updated on the progress of the scatterplot calculation After some minutes a pop up window should appear asking whether you want to view the scatterplot Hit Yes The usual IsoStar graphical interface will be opened if it is not already showing the calculated scatterplot of N H groups around methylcarbamoyl You can view and manipulate the scatterplot in the usual way see Tutorial 1 if you are not familiar with the operation of the IsoStar graphical interface Look at the files in your work directory You will find one called CONHMe istr This is the file containing the scatterplot Close the IsoStar graphical interface an
59. i are shown e Use of the hyperlinking facility shows that one of these occurs in the structure 1PPC a trypsin complex and involves the close approach of the backbone carboxylate of Asp189 SER190 7 5 Ethynyl O N S H e The CSD based plot of O N S H groups around ethynyl shows two interesting features e There are some hydrogen bonds to the acetylene pi system e There are several CH O interactions where the relatively acidic acetylenic hydrogen acts as an H bond donor to hydroxyl oxygen contacts shorter than the sum of van der Waals radii 0 2 A are shown TsoStar User Guide 49 e Again the hyperlink function can be used to inspect some of these For example one of the O H p1 H bonds occurs in the CSD structure BETXAZ 7 6 Aromatic I any C N O S or H e A particularly good way of spotting novel interactions is to look at scatterplots involving the contact group any C N O S or H e For example the CSD based plot for any C N O S or H around aromatic iodine shows several C I O contacts along the extension of the C I bond Lommerse et al J Am Chem Soc 118 3108 1996 Some of these are extremely short contacts less than the sum of van der Waals radii are shown below 50 TsoStar User Guide e Short highly directional sulfur oxygen and sulfur nitrogen contacts Burling amp Goldstein J Am Chem Soc 114 2313 1992 are found in the plot of any C N O S or H around sulfur heterocycle
60. iately plots placed in this TsoStar User Guide 53 54 directory are accessible to all IsoStar users within an organisation For more details click on Custom Plots at the bottom of the IsoStar central group menu and look at the ReadMe file TIsoStar User Guide 3 Nonbonded Searching Using ConQuest e Itis assumed that the reader has some knowledge of either ConQuest in particular how to build fragments and do simple substructure searches If information is required about these basic features the reader is referred to the relevant documentation www ccdc cam ac uk support documentation conquest e Alternatively the basics of searching using ConQuest are covered in tutorials provided at the end of the documentation Tutorial 5 covers nonbonded contact searching e The essential features of nonbonded searching in ConQuest will be illustrated with an example viz a very simple search for close nonbonded interactions between ketone oxygens and hydroxyl hydrogens see Section 3 1 e Some advanced search options are also covered see Section 3 2 3 1 A Simple Nonbonded Search e The first step is to draw the two groups using the ConQuest sketcher accessed from the Draw button in the main ConQuest interface File Edit Atoms Bonds 3D Options Help Click and drag to create Next Atom E a bond Sig Drag to an existing atom to make a connection Next Bond Single i DRAW EDIT ERASE ADD 3D CONTA
61. ically e Level use the up and down arrows to modify the contour levels e Color click on the coloured rectangles to change the contour colours e Show control the display of individual contours by switching the Show tickbox on or off Note that the display of contour surfaces can also be controlled using the Show All and Hide All buttons on the main graphical interface e To delete a contour first select it by clicking on the appropriate Contour entry in the Current Scatterplot table then hit Delete TsoStar User Guide e More surfaces at different contour levels can be added by repeating the above process e If you calculate a surface and nothing appears in the visualiser the probable reason is that the contour level is too high 3 4 2 Internal and External Scaling of Surfaces e When contouring a scatterplot there are two ways of defining the contour levels In nternal scaling the position of maximum contact atom density 1s arbitrarily assigned a contour level of 100 Every other density value is then scaled relative to this External scaling only available for CSD based scatterplots is more complicated An estimate 1s made of the average density of contact group atoms in all CSD structures containing both the central group and the contact group Then the actual density of contact group atoms at each grid point is divided by the average density This gives a figure that represents whether the density at that grid point is high
62. ide and then select the desired display property setting from the next menu Labelling Atoms Right click anywhere in the visualiser and select Labels from the resulting pull down menu then TsoStar User Guide 5 8 hit Show Labels to label the atoms currently selected see Section 5 4 If no atoms are selected then all atoms will be labelled You can also switch individual atom labels on and off by right clicking in the visualiser and selecting Picking Mode and then Pick Labels When the Pick Labels option 1s active left clicking on an atom will toggle its label on and off To remove all labels right click in the visualiser select Labels from the resulting pull down menu then hit Hide Labels The same pull down menu gives access to options for changing the size and colours of labels Measuring Distances Angles and Torsions To measure distances angles and torsions e Right click anywhere in the visualiser select either Measure or Picking Mode from the resulting pull down menu then select either Measure Distances Measure Angles or Measure Torsions or Pick Distances Pick Angles Pick Torsions Depending on which mode has been chosen you can then click on two three or four atoms respectively to measure a distance angle or torsion e You will remain in the chosen measurement mode so after measuring the first distance or angle or torsion you can continue measuring others To cancel the measurement mode right
63. in each grid volume This produces an empirical estimate of the density of contact group atoms at each of a regular matrix of grid points After application of a smoothing algorithm the densities are contoured at user specified levels and displayed as surfaces 3 4 1 Creating Surfaces e With a scatterplot loaded see Section 3 2 1 in the current visualiser see Section 5 1 click on the Create Edit button adjacent to Contour Surfaces 20 IsoStar User Guide Contour Surfaces Show All Hide All Create Edit e In the resulting dialogue window pick whether you want to use internal or external scaling see Section 3 4 2 e g Create New IsoaStar Contour Surfaces Internal Scaling 0 Level lt 100 C External Scaling 0 lt Level e Depending on which scatterplot is being contoured you may be given a choice of probes For example if the contact group is C O it will be possible to contour on the positions of the carbon or oxygen atoms Select the desired probe from the pull down menu e g Create New IsoStar Contour Surfaces Internal Scaling 0 lt Level lt 100 C External Scaling 0 lt Level Create E new surfaces using Probe carbon carbon Color om e Contour levels are provided however these may be user defined by typing new values into the Level boxes For internal scaling levels of between 20 to 90 are usually suitable for external scaling values between to 4 are often appropriate
64. inimum energy orientations for dimers are carried out using the ORIENT 3 0 Stone Popelier amp Wales ORIENT 3 0 A Program for the Calculation of Electrostatic Interactions between Molecules University of Cambridge Cambridge UK 1994 program This program can efficiently minimise the interaction energy of a given dimer starting orientation using a model intermolecular potential The model potential describes the long range electrostatic energy using the DMA multipoles by including all terms of the multipole expansion up to and including ae Repulsion and dispersion are described by an empirical isotropic exp 6 term This potential FIT Coombes Price Willcock amp Leslie J Phys Chem 100 7352 1996 reproduces a wide range of polar organic crystal structures and so will provide a reasonable first estimate of the intermolecular separations but ignores any anisotropy in the atomic repulsive wall The methodology described is adequate to compute long range interactions and therefore results in well orientated monomer positions for energy minimised dimers Since the method is fast it can be used to scan and minimise quickly the full intermolecular hypersurface Typically starting with 100 500 initial orientations the global and 5 20 local minima are obtained for any particular dimer e Intermolecular interaction energies are then calculated for selected minimum energy dimer orientations using the Intermolecular Perturbation Theor
65. is the current visualiser denoted by blue bars above and below the visualiser area For example in the screenshot below the current visualiser is the one on the left ocatterplot PDB charged carboxylic acid any polar 4 H 4 N 0 of 3 loyago ks f li p ri i Wale EESE Curentvisualiser e Ifyou click in a visualiser 1t becomes the current visualiser e The first scatterplot or theoretical energy minimum to be loaded into the IsoStar graphical interface will be placed in the left hand visualiser By default if a second scatterplot or theoretical energy minimum is loaded a dialogue will appear which allows you to specify which visualiser is to be used 1 e whether to overwrite the first scatterplot or use the empty visualiser E Overwrite Content Warning Loading this object will overwrite any content in the current visualiser Do not show this dialog again Cancel Load Load in Current Load in Other e If you switch on the Do not show this dialog again check box or if you select Options from the top level menu and disable Show Overwrite Visualiser Warning this dialogue will not be shown again and any further scatterplots or theoretical energy minima will be loaded into the current visualiser e When you hyperlink see Section 3 3 from a scatterplot the crystal structure will always be 28 TsoStar User Guide 5 2 5 3 loaded into the other visualiser i e the visualiser not containing t
66. less the user inputs alternative values e An alternative contact distance range can be input by clicking on Edit in the Non bonded contact definition window and activating the Distance range radio button The desired distance range can then be input e g 1f two distances are typed in 2 0 to 3 0 then all contacts in this range will be found 3 2 2 Searching for Contacts between Groups Rather than Atoms e In the above example a search was performed for contacts between the oxygen atoms of ketone groups and the hydrogen atoms of hydroxyl groups An alternative is to search for contacts between any atom of the ketone fragment and any atom of the hydroxyl group or between the centroids of the ketone and hydroxyl groups 56 TsoStar User Guide e This is done by clicking on the ADD 3D button clicking on each atom of the fragment in turn and then selecting the Define button next to Centroid on the Geometric Parameters window e Then do the same for the hydroxyl group Draw 1 New File Edit Atoms Bonds 3D Options Help DRAW EDIT ERASE ADD 3D CONTACT ile ke Ring aker Templates Select atoms to define 3D Parameters or Objects 2c ie E l0 x Defined nied CENT2FHBDG 20 Pareres oO ptio ns Delete Contacts Options To construct Parameter Object Select atoms main window or object list below Current Selection Valid Parameters Defined
67. ll down menu and then pick the required style Aromatic rings are displayed with a circle within them by default If this is not desired right click anywhere in the visualiser and select Display Aromatic Rings from the resulting menu Bond types i e single double triple etc are displayed by default If this is not desired right click anywhere in the visualiser and select Display Bond Types from the resulting menu By default atoms are coloured by element To change the colour used for a particular element right click in the visualiser pick Colours and then Element colours from the resulting menus and then hit the coloured box next to the element you want to change This displays a colour palette from which a new colour can be chosen Hitting Defaults at the bottom of the element colour dialogue will reset all element colours to their default settings If the colours of some atoms have been altered see Section 5 6 you can return to colouring by element by right clicking in the visualiser and picking Colours and then Colour by Element You can switch between the default black background and a blue gradient background by right clicking in the visualiser and hitting Draw Backdrop Setting Display Properties for Particular Atoms and Bonds To set the display properties of particular atoms select them see Section 5 4 right click in the visualiser select the appropriate option from the pull down menu Styles Colours Labels Show H
68. lots 12 Displaying Statistical Data 13 Displaying Theoretical Data 14 Looking at Closely Related Groups 15 PDB Plots of Ionisable Groups 16 Ligand and Protein Central Groups 8 The Example e This example should expand on what has been learned in tutorial 1 and introduce the user to the more advanced features of IsoStar 9 Selecting a Central Group e IsoStar contains information about a large number of chemical groups including terminal substituents acyclic linking groups and ring systems The major categories are listed on the left hand side of the IsoStar Web page TsoStar User Guide 9 92 gt IsoStar 2 1 Windows Internet Explorer j Y http fintranet ccde cam ac uk isostar jisostar_2 1 ntml isostar html File Edit View Favorites Tools Help Yr 2 v Search 2 A gt ant spy mal E Answers Dating GJ Mobie I Sign in w be IsoStar 2 1 IsoStar 2 1 Home Ligand Terminal C H only N C H only O C H only N O C H only Si containing P containing S containing Halo containing Acyclic links C H only N C H only O C H only N O C H only P containing S containing Ring systems Phenyls C H only Version 2 1 2009 Release N C H only O C H only N O C H only S containing Nucleic acid bases Solvates etc Inorganic Organic ane neg aaa naa Av etl PE Bel PES USS ES BEE ay Welcome to soStar g e Do not worry at this stage about the distinction bet
69. lp The context dependent help facility can be accessed by clicking with the right hand mouse button on any option in the IsoGen interface This opens a pop up which gives a brief reminder of the function of the chosen option e About IsoGen displays a pop up window which explains what IsoGen is who distributes it and who should be contacted in case of difficulty TsoStar User Guide 6l 5 Isogen Expert Features oD symmetry Atom ty pes Slider atoms Omit atoms Flexible atoms Contour probes e A number of options are available under the top level Expert menu option Their functions and use are described in the following sections e 3D Symmetry see Section 5 1 e Atom types see Section 5 2 e Slider atoms see Section 5 3 e Omit atoms see Section 5 4 e Flexible atoms see Section 5 5 e Contour probes see Section 5 6 e Use dummy atoms in fitting see Section 5 7 5 1 3D symmetry e The 3D symmetry of the central group is used by the program for two reasons e to calculate an average geometry for the central group 2 to reduce the scatterplot so that all contact groups lie in one asymmetric unit of the central group s point group This is illustrated by the following raw and symmetry reduced plots in the latter all contacts have been reflected into one quadrant since the central group has mm symmetry 62 TsoStar User Guide e The 3D symmetry menu option offers three possible selections De
70. luded in IsoStar intramolecular contacts are not considered e In both the CSD and PDB sections of IsoStar nonbonded contacts are included up to a maximum distance of V Tol where V is the sum of the van der Waals radii of the atoms involved and Tol is a distance tolerance Currently a distance tolerance of 0 5 A is used Van der Waals radii are taken from Bondi J Phys Chem 68 441 1964 H 1 20 C 1 70 N 1 55 O 1 52 F 1 47 Si 2 10 P 1 80 1 80 Cl 1 75 A Br 1 85 I 1 98 6 1 4 Substructure Search Details e A central group see Section 1 1 in IsoStar usually contains two types of atoms target atoms and non target atoms Non target atoms are there to help define the chemical environment of the group e g the carbon atom in C sp NH but are not included in nonbonded searches In other words only nonbonded contacts to target atoms are shown on IsoStar scatterplots For example scatterplots for C sp NH only show contacts to the nitrogen and hydrogen atoms e Many of the central groups are ring systems e g thiazole In any particular CSD or PDB crystal structure the ring is almost certain to be substituted For example one of the structures containing thiazole in the CSD BAWKEP10 is substituted at the 2 position TsoStar User Guide 33 e In IsoStar contacts to ring substituents e g the amino group above are not shown However contacts to
71. mm and 3m symmetry respectively 3 2 7 Customising a Scatterplot Display e Clicking anywhere in a visualiser with the right mouse button will display a menu which allows you to alter the visual appearance e g atom colours and styles of a scatterplot see Section 5 3 3 Hyperlinking from a Scatterplot to a Crystal Structure e Hyperlinking is only operational when the current visualiser see Section 5 1 contains a I8 TsoStar User Guide scatterplot and when the Hyperlink check box has been switched on Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab e In this situation clicking on any contact group atom in the scatterplot will cause the crystal structure from which that contact was taken to be displayed in the other visualiser window e g Contour Surfaces Show All Hide All Create Edit Scatterplot Symmetry Expand Contract V Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab Scatterplot CSD nitro cationic R3NH Current visualiser Filename C DOCUME ROBERT I LOCALS 1 Temp csd0042_4 1 istr compressed e The contact that was selected from the scatterplot will be highlighted in green this makes it easier to select contacts from a cluster without accidentally picking the same one twice The highlighted contacts can be set to their original colours by right clicking in the scatterplot visualiser selecting Colours from the resulting men
72. nctional groups and contacts to similar functional groups in the higher precision small molecule CSD structures In general tighter chemical constraints have been used in selecting the CSD structures that contribute to the plots in the Protein section For example any CSD structure containing a C CONH b group will contribute to the carbamoyl CSD based scatterplots in the Ligand section However only structures containing the more precisely defined substructure CH5 CONH will contribute to the Protein carbamoyl CSD plots 2 4 Conformationally Flexible Central Groups e When a central group see Section 2 1 adopts two or more clearly distinct conformations each is included in IsoStar as a separate entry e g carbamate trans cis and carbamate trans trans carbamate trans cis carbamate trans trans e An exception may be made if one of the conformers is much more common than the other s in which case only the major conformer may be included e Sometimes the division between two conformers may not be distinct 1 e some structures may lie half way between two minimum energy geometries In this case an arbitrary criterion 1s used For example anilines are considered to be planar if the sum of the three bond angles at nitrogen exceeds 352 5 otherwise they are considered to be pyramidal The criterion of 352 5 is used throughout IsoStar to define planar and pyramidal nitrogen e Some central groups have ill defined conformation
73. ne Theoretical Chemistry Department University of Cambridge UK and Sally Price Department of Chemistry University College London who provided software for and advice about molecular orbital calculations e Gerhard Klebe University of Marburg Germany Stefan Brode and Hugo Kubinyi both of BASF AG who provided benchmark molecular orbital data e David Watson CCDC and Manfred Hendlich University of Marburg for providing information about the chemical connectivities of protein ligands in the PDB TsoStar User Guide 79 80 Appendix 5 IsoStar Tutorials Tutorial 1 Manipulating IsoStar Scatterplots Tutorial 2 Navigating the IsoStar Web Pages Tutorial 3 Calculating Customised Scatterplots Using IsoGen UNIX only TIsoStar User Guide Tutorial 1 Manipulating IsoStar Scatterplots The Example Getting Started Exploring the Visualiser Interface and Interacting with a CSD Plot Hyperlink to a CSD Structure Generation of Contoured Density Surfaces Inspecting a PDB Scatterplot Hyperlink to a PDB Structure Inspection of a Theoretical Plot SAAN WN 1 The Example e Isostar is a library of information about the nonbonded interactions formed by a wide variety of chemical groups It is based on data from the Cambridge Structural Database CSD the Protein Data Bank PDB and molecular orbital calculations It is hoped that this example will introduce the user to IsoStar and its sources of data and demonstrate some of the ba
74. of Initial Model e An approximate average geometry for the central group is derived as follows The first central group fragment in the ConQuest coordinate file is used as a mode onto which the second fragment is fitted The average geometry of these two fragments is computed The third fragment is then fitted onto this average The average geometry is again updated The process is repeated until all fragments in the ConQuest file have been fitted giving a final average model for use in the next step e Problems may occur with central groups that contain conformationally flexible moieties e g rotationally mobile methyl groups In such cases the fitting procedure may result in artificially shortened average bond distances 6 4 Superposition e A second pass through the data is carried out in which each fragment is fitted onto the average model obtained above Standard methodology is used for the fitting Mietzner 1997 Fragments are rejected i e eliminated completely from the scatterplot calculation if they fit onto the average geometry with an RMS deviation exceeding 0 5 A Fragments are also rejected if after fitting they contain any individual atom that lies more than 1 0 A from the corresponding atom in the average model Currently these tolerances cannot be changed by the user e In cases where topologically equivalent atoms have been detected torsion angles are used to TsoStar User Guide 73 6 5 resolve which atoms match
75. ogen X H where X is N O or S e Hydrophobic groups e g methyl phenyl e N H hydrogen bond donor groups e g amide NH ammonium NH e Various types of O H groups e g alcohol OH water e Other groups containing N and or O Most of these are H bond acceptor groups e g cyano carbonyl O nitro e Various types of sulfur e g thioether thiocarbonyl e Groups containing halogen atoms e g C F chloride e Groups found in amino acids e g imidazole guanidinium Ligand and Protein Classification of Central Groups Central groups see Section 2 1 in IsoStar are divided into two main sections Ligand and Protein The distinction is important for the PDB part of the library and is best explained by an example Consider the central group carbamoyl CONH 3 This appears in both the Ligand and Protein sections However the two sets of PDB scatterplots see Section 1 1 are not identical This is because the plots in the Ligand section show contacts to carbamoyl groups on ligand molecules Conversely the plots in the Protein section show contacts to the carbamoyl groups of asparagine and glutamine side chains in proteins Strictly speaking all CSD based scatterplots should be under the Ligand section of IsoStar since the CSD contains only small molecules not proteins However CSD data are included in the Protein section as well so that comparisons can easily be made between non bonded contacts to TsoStar User Guide protein fu
76. on is switched on these atoms are included in the least squares superposition of central group fragments They can be omitted from the fitting calculation by switching the button off TsoStar User Guide 71 6 IsoGen Theory The Scatterplot Calculation Procedure 6 1 Summary e Before IsoGen can be run the user must have performed a ConQuest nonbonded search and save 6 2 2 a number of output files namely the tab fgd and fgn using the Export Parameters and Data option from the File menu and the cor using the COORD CSD Coordinate file file type from the Export Entries As option found under File The cor file contains the fractional crystallographic coordinates of each nonbonded contact found in the ConQuest search together with some symmetry information From this starting point the key steps in producing a scatterplot are e Definition of program settings via the IsoGen graphical user interface e Reading of data from ConQuest output files e Detection of any topological symmetry in the central group the group which will be overlaid e Pass 1 Derivation of an initial average geometry for the central group e Pass 2 Superposition of all central group fragments in the ConQuest output file onto the average group derived in the preceding step Conformational outliers are eliminated at this stage Torsion angles are used to resolve ambiguities due to topological symmetry e Derivation of 3D molecular symmetry if
77. ons tick box This will instruct ConQuest to standardise all N H bond lengths to 1 009 A before searching for intermolecular contacts This is an empirical way of correcting for the fact that N H bond distances determined by X ray diffraction are usually too short e Now hit Start Search to set the search running You will be taken to View Results window where a scrolling list of hits containing the substructure and contact 1s displayed 5 Calculating a Scatterplot e To calculate a scatterplot from ConQuest data you will need to save a number of files from the search you have just run So create a new directory for these files in your home area e In ConQuest export the 3D parameters and data by clicking on File and selecting Export Parameters and Data from the pull down menu Ensure that the Include Defined Parameters tick box is activated and click Save Browse to the directory you just created and save the files as e g CONHMe tab e The folder will contain three files with the extensions tab fgn and fed e Now export the fractional fragments by clicking on File and selecting Export Entries As Select COORD CSD Coordinate file from the Select file type pull down menu and ensure that the All selected entries Fractional and Hit Fragment Only radio buttons are activated This is particularly important as the Isostar plot will not be successfully generated if these buttons are not activated Browse to the directory you created and save t
78. onto which For example the Z hydrogen of vinyl H2 in the diagram above can be distinguished from the E hydrogen H71 because the torsion angle C C C H2 will be close to zero whilst the C C C H1 torsion will be approximately 180 degrees When topologically equivalent atoms cannot be distinguished on the basis of torsion angles they are assumed to be symmetrically related in 3D Derivation of 3D Symmetry The 3D symmetry is derived in a series of steps First the 2D connectivity Morgan algorithm scores are used to deduce the maximum order of symmetry that the central group could possibly have assuming that the group is not completely linear If there are no topologically equivalent atoms the central group can have nothing higher than mirror symmetry with all atoms lying in the mirror plane The number of topologically equivalent atoms in a group defines the maximum order of symmetry by which these atoms can be related and so defines the maximum order for searching For lower symmetry central groups the first step in locating symmetry is to transform the group to its moments of inertia In higher symmetry systems the moments of inertia are not defined uniquely and so are inappropriate This can be visualised by considering elliptical solids An ellipse that has three radii of differing lengths has mmm symmetry the directions of the 3 principal axes are unique Next consider the case of an ellipse with 2 radii of equal length and one diff
79. ot that shows the distribution of C N O S and H atoms around carboxylate anions in the CSD The plot may take some time to load If the central group is unclear its style can be changed e g from wireframe to capped stick by selecting each atom and picking the relevant option from the right click menu Return to tutorial 1 1f you are unsure how to do this In the plot polar hydrogen atoms OH NH and SH hydrogens are coloured pale green other hydrogens virtually all of which will be from CH groups are coloured white Carbons are grey oxygens red and nitrogens blue TsoStar User Guide e You can see the clusters of O N and H atoms in the carboxylate oxygen lone pair directions corresponding to OH carboxylate and NH carboxylate H bonds e Look back at the number you hit in the Web browser to display this scatterplot This is the number of contact groups in the plot Because carboxylate is a common group the scatterplot would contain tens of thousands of atoms 1f it included all of the contacts in the CSD This would make it very slow to load and manipulate In setting up IsoStar therefore the plot has been limited to a total of about 5 000 atoms This was done by taking a pseudo random subset of the available CSD data e Close the IsoStar graphical interface and return to the contact table in the Web browser e To investigate H bond geometries more closely click on the number in the row marked any polar X H and the column head
80. oxylate contacts are very common and therefore by implication energetically favourable e Click the browser Back button to return to the contact group table 13 Displaying Theoretical Data e You will notice some round gold coloured buttons in the contact group table These indicate that results from molecular orbital calculations are available e Go to the section of the contact group table headed O H General C H only N H O H Other N or O Sulfur Halo halide Amino acid O H Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any OH 2480 4888 amp alcohol OH 1661 1129 gt phenol OH 352 794 water 1585 4767 gt This search has not been done TsoStar User Guide 97 e Click on the gold button in the row marked alcohol OH and the column headed Theory This displays a summary of calculations performed on the model system acetate methanol The table shows the interaction energies of the most important minima broken down into different physical contributions The total interaction energies in the right most column are very large This is because the calculations are on a gas phase model system and one of the interacting species 1s charged e Click on the number 1 in the column headed Minima This will display the geometry of the lowest energy minimum e Now click on the number 2 to display the second lowes
81. quencies and directionalities of intermolecular contacts The above example shows that OH groups the contact group in this case strongly hydrogen bond to the carbonyl oxygen of esters the central group but not to the other oxygen atom e Each scatterplot can be converted into a contoured density surface which is often useful for TsoStar User Guide 3 highlighting geometrical preferences e g e The molecular orbital data provide information about interaction energies and theoretical in vacuo minimum energy geometries e g e IsoStar also contains statistical data on the frequencies with which various interactions occur in crystal structures 1 2 Overview of the IsoStar Interface e IsoStar uses a standard web browser to allow selection of the interaction of interest from a pre computed library of several hundred central groups see Section 1 1 and about fifty contact groups e Information for each central group is presented in a table which provides links to scatterplots tabulated MO results and statistical data e Scatterplots can be displayed in a graphical interface which allows 4 TsoStar User Guide e customisable 3D visualisation of the scatterplot see Section 5 e control by distance over which contacts are displayed see Section 3 2 2 e presentation of the scatterplot as a contoured density surface see Section 3 4 e hyperlinking to CSD or PDB entries that contribute to the scatterplot see Section 3 3
82. r S 2465 386 This search has not been done e Clicking on the adjacent number 3 6 will show the scatterplot of nitro H X contacts observed in protein ligand complexes from the PDB e Ifa cell in the CSD or PDB columns contains an asterisk 1t means that no scatterplot has been generated for that particular contact normally because there are no experimental data e The selected scatterplot is loaded into one of the two 3D visualisers in the IsoStar graphical interface 14 TsoStar User Guide E Isostar File Options Help Previous Files Show contacts in van der Waals vyd corrected distance range Scatterplots Hyperlinks Theoretical Energy Minima Lower Limit AONAONOONONOONOONOONOONOONOONONOONOONOONOONOONONOONOONOONOONOONONOONOONOONOMN AMNEM Le CSD jnitro any polar X H lt N O or S Upper Limit CUP ECEC UP OE CADE CEC OPE E CUE C UD OP EEE OECD CECT EEE AE EEC nin ey 0 5 Hide All Overview a vaii Overlaps Show All Contour Surfaces Show All Hide All Create Edit Scatterplot Symmetry Expand Contract Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab Scatterplot CSD nitro any polar X H lt N PDB Current visualiser Filename C DOCUME ROBERT 1 LOCALS 1 Temp csd0042_02 istr compressed e The graphical interface has options for controlling which visualiser is used when further scatterplots are loaded see
83. r User Guide E CSD charged carboxylic acid any GH Contact Group Information Identifier Distance A 4 Cumulative Distribution BOOVEIO 1 1 27 wt SOO ae ICBE 1 27 2000 ZACE 1 27 1500 1000 ees 500 Al 0 1 5 0 5 0 0 5 OKIOE 1 26 HOCMTO4 1 26 MOK 1 26 ra ee van der Waals corrected distance Angstrom Frequency Distribution IFSUA 21 25 ABPEP 1 25 g ISAR 1 25 LYVOM 1 25 EVMEQ 1 25 20 j ij j EJMOY 1 25 m O f il 1 1 25 3 x 1 5 0 5 0 0 5 van der Waals corrected distance Angstrom at aa Li mI 1 t m ht 5 J gt JE i TA e Clicking on a refcode e g TULWOM in the dentifier list selects the contact group in the scatterplot e The Cumulative Distribution provided in the top right of the dialogue graphs the number of contacts versus the contact distance the contact distance is expressed relative to the sum of van der Waals radii of the contact atoms e The Frequency Distribution shows the number of contacts at each contact distance 3 2 4 Expanding and Contracting Scatterplot Symmetry e In IsoStar scatterplots are symmetrised 1 e all the contacts are reflected into an area of space based on the symmetry of the central group e g only one quarter of an aromatic iodine group with its contact group is shown since the central group has two planes of mirror symmetry whereas half of all contacts to an azide
84. related thermolysin complexes 6 2 5 Biases in Central Group Geometries e The central group geometry in a scatterplot is obtained by translating and rotating individual structures so that the central group moiety is in a standard position and orientation A mean geometry is then obtained by averaging in Cartesian coordinate space For a rigid group this produces an average geometry very close to that which would be obtained if the averaging were done in internal coordinate bond length bond angle torsion angle space For more flexible groups however the averaging of Cartesian coordinates results in a slight shrinkage of the bond lengths of the group similar to that produced by the librational effect in crystallography This has an insignificant effect on the IsoStar scatterplots However the central group geometries in IsoStar should not be taken as the statistically optimum geometries of these groups 6 3 Details of Molecular Orbital Methodology e The ab initio package CADPAC 6 0 Amos CADPAC6 0 The Cambridge Analytical Derivatives Package University of Cambridge Cambridge UK 1996 is used to calculate the wave functions of individual monomers and to optimise their geometries at the 6 31G basis set level The charge distributions are corrected for electron correlation by the MP2 method A distributed multipole analysis DMA is then carried out on the wavefunctions to obtain atomic multipoles up to hexadecapole e Calculations of m
85. rive Define and Do not use The default option is Derive This makes the program deduce the 3D symmetry automatically from the ConQuest output a procedure which usually gives satisfactory results Do not use instructs the program to produce a raw plot 1 e with no determination of 3D symmetry As the name suggests the remaining option Define allows the user to specify explicitly a particular symmetry Selecting this option causes the following window to be opened Set Symmetry ok vewsearen cancer i TsoStar User Guide 63 e This window enables the user to specify explicitly up to three symmetry elements The required element 1s selected by clicking on one of the left hand buttons and choosing from the resulting list of elements The direction in which the symmetry element lies is then typed into the corresponding white box Directions are defined using central group atom atom vectors or centroid atom vectors see Appendix Vector Direction Syntax page 76 Note that the central group atom numbers may be displayed by clicking on View Search It is possible to specify sums and cross products of bond vectors as well as bond vectors themselves this is most easily illustrated with examples e Example 1 Ketone see Section 5 1 1 e Example 2 Benzene see Section 5 1 2 e Example 3 Sulfate see Section 5 1 3 5 1 1 Defining Symmetry Example 1 Ketone e T
86. rm in whole or in part to any third party e Software or data derived from or developed using the CSD System may not be distributed without prior written approval of the CCDC Such prior approval is also needed for joint projects between academic and for profit organisations involving use of the CSD System e The CSD System and its Components may be used for scientific research including the design of novel compounds Results may be published in the scientific literature but each such publication must include an appropriate citation as indicated in the Schedule to the Licence of Access Agreement and on the CCDC website e No representations warranties or liabilities are expressed or implied in the supply of the CSD System or its Components by CCDC its servants or agents except where such exclusion or limitation is prohibited void or unenforceable under governing law Licences may be obtained from Cambridge Crystallographic Data Centre 12 Union Road Cambridge CB2 1EZ United Kingdom Web http www ccdc cam ac uk Telephone 44 1223 336408 Email admin ccdc cam ac uk UNITY is a product of Tripos L P and MDL 1s a registered trademark of Elsevier MDL Chapter 1 IsoStar User Guide 1 NTA bh WN Introduction Organisation and Navigation of IsoStar Data Viewing and Using Scatterplots Viewing Other Data Using the 3D Visualiser Experimental Details and Limitations Example Results Chapter 2 IsoGen User Guide 1
87. s For example the C S S C torsion angle of acyclic disulfide linkages is usually about 85 but can be as low as 50 or as high as 120 In such a case an average geometry is calculated and individual structures are omitted if they do not overlay well enough on this average specifically all non hydrogen atoms must fit to within 1 A and the rms deviation must be less than 0 5 A e In some cases a central group adopts a well defined conformation but contains a terminal group such as methyl or hydroxyl that shows a great deal of rotational flexibility In this case the H atoms may be omitted entirely or displayed as shown in the scatterplot below TsoStar User Guide 7 2 5 Central Groups with Ambiguous Protonation States e Ifa central group see Section 2 1 can exist in more than one conformer or more than one ionisation or tautomeric state the various forms will be included separately e g C OPO3 and C OPO3H are separate entries in the library 2 6 Accessing Data for a Particular Central Group e To obtain information for a particular central group see Section 2 1 open the IsoStar home page in a web browser The available types of central groups are listed on the left hand side of this page divided firstly by whether the central group 1s in the Ligand or Protein section see Section 2 3 then by whether the group is a terminal substituent linking group ring system or solvate molecule and then by the element s it contains
88. s contacts shorter than the sum of van der Waals radii 0 3 A are shown below TsoStar User Guide 5 52 Nn bk OQ N IsoGen Quick Summary Introduction to IsoGen Nonbonded Searching Using ConQuest Isogen Basic Features Isogen Expert Features IsoGen Theory The Scatterplot Calculation Procedure TIsoStar User Guide 1 IsoGen Quick Summary e To generate a scatterplot from the CSD the following simple recipe will usually suffice e Perform a ConQuest nonbonded search for the contact of interest e Save the 3D parameters using the Export Parameters and Data option from the File menu This will result in 3 files lt search name gt tab lt search_ name gt fgn and lt search name gt fed e Then you will need to save the fractional fragments using the COORD CSD Coordinate file option in the Export Entries As part of the File menu e After the search has run browse to the directory where the files are stored and type LSOGer Search name gt e at the Unix prompt where lt search name gt is the name used for the ConQuest search e Pick Run from the IsoGen top level menu If all is well the scatterplot will be calculated and saved as a file with the extension lt search_ name gt istr 2 Introduction to IsoGen e IsoGen is the program that was used to calculate all the scatterplots in the IsoStar database It is released to users so that they can calculate their own customised scatterplots e g for groups
89. s and the corresponding dimer orientations are stored in the library Limitations of Molecular Orbital Results Calculations are usually performed on the simplest possible model compounds Consequently steric effects are usually less important in the model systems than in the many highly substituted molecules found in the CSD and PDB As with all molecular orbital calculations the accuracy of the results is limited by the basis set used and the methodology employed However comparisons between the method used in IsoStar and extended basis set triple zeta plus polarisation supermolecule calculations show an encouraging degree of concordance Brode amp Lommerse unpublished work 1996 There is no way of ensuring that all minima in a potential energy surface are found For any given model system therefore it is always possible that some important energy minima have been missed In cases where this seems particularly likely an appropriate warning message has been included in IsoStar By far the biggest limitation in the theoretical calculations is that they model an in vacuo situation They are therefore unsuitable for indicating the gain or loss of free energy that occurs when a group 1s taken out of solution to form a nonbonded interaction to a protein Also weakly attractive interactions that occur in vacuo are much less likely to be found in condensed phases where competing interactions are possible For example the calculated minimum
90. s dialogue it is possible to modify contour levels using the up and down arrows adjacent to the Level value and the contour colour Whether a contour is displayed or not can also be controlled using the Show tickboxes e Close the Contour Surfaces window using the Close button and return to the main IsoStar interface e Both the scatterplot and the contour plot are present in the 3D view so remove the scatterplot by activating the Hide All tick box N e You can also experiment with External Scaling settings e Hide the existing contours by clicking on the Hide All button then open the Contour Surfaces window by clicking on the Create Edit button e In the Contour Surfaces window activate the External Scaling radio button and ensure the probe is still set to Hydrogen e As before select new contour colours if desired but leave the Level settings as they are Once you are happy with the settings hit Create e The new surface is scaled relative to the density of hydroxyl hydrogens that would be expected by chance given the number of these atoms that occur in CSD structures So the yellow contour shows where the density of hydroxyl hydrogens is two times as high as would be expected by chance The red contour corresponds to a density expected by chance and so on e Close the Contour Surfaces window by hitting the Close button then close the IsoStar graphical interface 6 Inspecting a PDB Scatterplot e Click here to display
91. screen by keeping the left hand mouse button and the Shift key pressed down Translate by moving the cursor in the visualiser with the centre mouse button depressed requires 3 button mouse Alternatively use the left hand button with the Control key pressed down Zoom in or out by moving the cursor up and down in the visualiser while keeping the right hand mouse button pressed down TsoStar User Guide 29 5 4 5 5 5 6 5 7 30 Selecting and Deselecting Atoms Selection of atoms and molecules is useful for changing properties such as display style Atoms may be selected or deselected in several ways e Right click anywhere in the visualiser atom bond or background and choose Selection from the resulting menu Various options for selecting atoms can then be chosen from the resulting menus e Right click in the visualiser select Picking Mode from the resulting menu and set the mode to Default Picking Mode Click on individual atoms with the left mouse button to select them Once selected an atom can be deselected by clicking on it again e When the picking mode is set to Default Picking Mode all atoms become deselected if you left click anywhere in the display area background Setting Global Display Options A choice of four display styles is offered wireframe capped stick ball and stick and space filling To change the display style globally right click anywhere in the visualiser Select Styles from the resulting pu
92. sic features of the program 2 Getting Started e To start the tutorial click here If IsoStar is installed correctly the scatterplot will be displayed in one of the two visualisers in the IsoStar graphical interface Move the window to a convenient location on the screen e The visualiser window contains a scatterplot showing the distribution of OH groups around aliphatic ethers in crystal structures taken from the CSD The plot was prepared by finding all OH ether contacts in the CSD that lie within van der Waals radii 0 5A and overlaying them so that the ether portions were exactly superimposed Because the ether group 1s symmetrical all OH groups have been reflected into one quadrant TsoStar User Guide S1 Gi IsoStar Bel x File Options Help Previous Files Show contacts in van der Waals vdvV corrected distance range Scatterplots Hyperlinks Theoretical Energy Minima Lower Limit CUP CECE CC UU URGE CE OUD CECE CDE OE CECE DOE DE CEE EEE CEE PEE EEE EEE EEE EE EEE EEA EPEAT EEE AE ECE E AE EEE EAE ESET AT AEE OOOD ON Eee 1 21 CSD aliphatic aliphatic ether any OH CUECECE CECE CECE CECE CECE AERC AD AE AD ADAP EDAD ADEE CECE CECE CECE CECE CECE OEE Upper Limit A EE N 0 5 PDB Hide All Overview vd Overlaps Show All Contour Surfaces Show All Hide All Create Edit Scatterplot Symmetry Expand Contract Hyperlink Clicking on scatterplot atoms adds parent structure to Hyperlinks tab Scat
93. sity surface e The corresponding PDB based scatterplot below shows similar features contacts are shorter than the sum of van der Waals radii 0 5 A Now however only the oxygen atoms of the OH groups are shown as H atoms are rarely located in protein crystal structures For the same 44 TsoStar User Guide reason we cannot be sure that all the contacts on this plot are to ionised rather than neutral acid groups e Theoretical IMPT calculations on the model system acetate methanol also support the conclusion that hydrogen bonds to carboxylates form preferentially along the lone pair directions The two lowest minima occur in these positions 7 2 Aliphatic ether OH e Not all H bond acceptors show lone pair directionality For example the CSD based plot of OH around aliphatic ether shows no particular clustering in the lone pair tetrahedral directions contacts shorter than the sum of van der Waals radii 0 5 A TsoStar User Guide 45 e The CSD based plot of OH around aliphatic ester has several interesting features contacts shorter than the sum of van der Waals radii 0 7 e First the carbonyl oxygen accepts H bonds frequently but the C O C ester oxygen almost never accepts Secondly one of the carbonyl lone pair directions is preferred to the other this becomes clear if the scatterplot is contoured e The first observation is confirmed by IMPT calculations on the model system methyl acetate m
94. t Brackets are not currently supported so if one specifies a vector as Lee A gc ae age the program will interpret this as the cross product of the sum of 2 4 and 2 5 with 2 3 i e a b c where a b and c are the three vectors Conversely Dye Se Lig Fe 2S would be interpreted as the cross product of 2 4 and 2 5 summed with 2 3 i e a b c TsoStar User Guide Appendix 2 Algorithm for Identifying Topological Equivalence e The algorithm used is a modification of the Morgan algorithm and is best explained with an example H2 Let us label the carbons C Cy and Cy 779 and the hydrogens H H and Hy Obviously only atoms of the same elemental type can be topologically equivalent Thus it is immediately clear that the carbon atoms can be separated from the hydrogen atoms The algorithm proceeds by analysing the extended connectivity in the following way A score is assigned to each atom Initially the scores are computed by counting the number of bonds formed by each atom 1 e C 1 Cy 3 and CHp 3 This tells us that C is unique hence amongst the carbons only Cy and Cy 479 can possibly be topologically equivalent All the hydrogens have a score i e sum connectivity of 1 e In the second iteration the new score of each atom is calculated by summing the first iteration scores of all the atoms to which it is bonded CH gets a score of 1 C 1 H 3 Cym 5 Crip gets a score of 3 Cp 1 H 1 H2
95. t minimum The theoretical results support the crystallographic data in suggesting that the H bonds prefer to form along lone pair directions e Close the IsoStar graphical interface and return to the Web browser window 14 Looking at Closely Related Groups e So far we have looked at carboxylate anions IsoStar also contains data for unionised carboxylic 9S TsoStar User Guide acids which may exist in two different conformations O C O H cis or trans These are shown at the top of the Web page e Click on the hyperlink uncharged carboxylic acid cis to display the contact group table for this group uncharged carboxylic acid cls e Go to the section of the contact group table headed O H and click on the number in the row marked water and the column headed CSD You will get a scatterplot showing the CSD distribution of water molecules around cis carboxylic acid groups It shows that the acid carbonyl oxygen is a much better H bond acceptor than the acid OH oxygen which accepts only rarely Note the cluster of H bond acceptor water molecules near the acid OH hydrogen e Close the IsoStar graphical interface and return to the Web browser 15 PDB Plots of Ionisable Groups e Now for an important point about the PDB based scatterplots of acidic and basic groups Make a note of any number in the PDB column of the contact group table and compare it with the corresponding numbers for charged carboxylic acid and uncharged carboxylic
96. t not in the PDB Ch CH a al i D gt e Consequently IsoStar contains separate CSD based plots for different tautomers but only a single combined PDB based plot For this and other reasons the symmetry of CSD and PDB scatterplots sometimes differs e Imidazole as in histidine illustrates several of the problems with PDB based scatterplots There is a tautomeric ambiguity 4 yl or 5 yl an uncertainty in ionisation state neutral or cationic and at poor resolution the crystallographer may have had difficulty in distinguishing ring nitrogens from ring carbons This makes PDB based plots for this ring particularly difficult to interpret For example in the plot of OH around imidazole from histidine sidechains it is not clear which OH groups are donating to ring nitrogens and which are accepting from them There are also some short contacts on this plot between hydroxyl oxygens and the imidazole C2 atom these may be CH O hydrogen bonds or may be errors resulting from the imidazole ring being fitted to the electron density the wrong way round The contacts shown in the scatterplot below are shorter than the sum of van der Waals radii 0 2 A TsoStar User Guide 37 6 2 3 Biases in CSD Based Scatterplots e There are two reasons why a CSD based scatterplot might be misleading because of a common substitution pattern in the database or because of a frequent crystal packing motif The first is illustrated by the distribution of car
97. terplot CSD aliphatic aliphatic ether any OH Current visualiser Filename C DOCUME 1 ROBERT 1 LOCALS 1 Temp getting_started_csd_plot_1istr e Contact groups can be viewed for the remaining quandrants using the Expand button next to Scatterplot Symmetry If you do expand the symmetry return the scatterplot to its initial state using the Contract button e The scatterplot can be rotated with the left mouse button and translated with the middle mouse button or using the left mouse button whilst pressing on the Control key on your keyboard The plot can be zoomed in or out using the right mouse button e Inspect the scatterplot It shows that OH groups tend to lie quite close to the plane bisecting the C O C angle Within that plane they can adopt a variety of positions 3 Exploring the Visualiser Interface and Interacting with a CSD Plot e A number of options for modifying the scatterplot can be displayed by right clicking in the 3D display e Select the three atoms that make up the ether moiety 1 e COC and right click anywhere in the 3D display From the resultant menu select Styles then Capped Sticks The ether central group will now be displayed in the capped stick style while the OH contact groups are displayed in the 82 TsoStar User Guide default wireframe e The scatterplot currently displays all OH ether contacts in which the ether oxygen is within a distance of V 0 5 A to either the hydroxyl O or H atom
98. to remove them When the pop up window appears asking if you want to view the scatterplot select Yes You should now see the various orientations of the methyl group shown explicitly something like this TsoStar User Guide 109 e Close the IsoStar graphical interface and hit Ok in the IsoGen text window 7 Omitting Atoms e Another thing we can do is omit conformationally variable atoms altogether First we need to undo our previous Flexible atoms definition Select Expert from the IsoGen top level menu and Flexible atoms from the sub menu Delete the numbers from the white box in the resulting pop up window and hit Ok e Now select the IsoGen option Expert and then Omit atoms The search fragments are again displayed in a pop up window Pick the methyl group H atoms and hit Ok e Hit Run in the IsoGen window to re compute the scatterplot This time as you will see the hydrogens of the methyl group are not displayed at all e Finally remove the windows that remain from the tutorial The main IsoGen window can be removed by hitting File and then Exit IsoGen 110 TsoStar User Guide This finishes the tutorial which has demonstrated how to produce scatterplots from the CSD by performing nonbonded searching in ConQuest and then post processing the output with IsoGen Scatterplots produced in this way may be made accessible from the IsoStar web page by putting them in the Custom Plots area TsoStar User Guide 11l 112 TIso
99. u and then Colour by Element from the next menu e In the crystal structure display the atoms involved in the contact are highlighted by being displayed in ball and stick mode e The database identifier of the structure that has been loaded will be displayed in the Hyperlinks area top left of interface either in the CSD or PDB box depending on which database the structure came from TsoStar User Guide 19 Previous Files Scatterplots Hyperlinks Theoretical Energy Minima Identifier Contact number Scatterplot e Hitting the Info button next to the structure identifier will display further information e g literature reference about the structure 3 4 Displaying Contoured Density Surfaces e Scatterplots can be converted to contoured density surfaces For example the scatterplot of carbonyl groups around iodo substituents see Section 3 1 can be displayed as shown below This surface highlights the regions in space where the carbonyl oxygen atoms accumulate e Here the contour colours have been chosen so that red denotes regions preferred by the O atoms and yellow denotes the most preferred region In a similar way a surface could be produced showing where the carbonyl carbon atoms tend to be e The surfaces are calculated by dividing the space around the central group into a regular grid and counting the observed number of contact group atoms carbonyl oxygens or carbons in this case
100. uctures means that there are no PDB plots for the central group water since the water molecules cannot be overlaid 6 2 Scatterplot Problems TsoStar User Guide 35 6 2 1 Experimental Errors e Although CSD crystal structures are generally of high accuracy the database inevitably contains some experimental errors These are usually in the placement of hydrogen atoms For example the CSD crystal structure ADGLAL10 shows an impossibly short H H contact of 1 14 A Clearly a hydroxyl or water H atom has been misplaced in this structure e Errors in PDB crystal structures are much more common because of their poorer resolution However the increasing use of refinement programs such as X PLOR has tended to reduce the number of impossibly short contacts in protein crystal structures by including an empirical potential energy term in the minimisation function While reducing the number of obvious errors this introduces a subjective bias into the results since they reflect to some degree the empirical parameters used in the force field Experimental errors can be very noticeable in scatterplots because they often appear as outliers Some obvious errors have been removed from the IsoStar library specifically any contact shorter than V 1 3 A has been removed where V is the sum of the van der Waals radii of the atoms involved 6 2 2 Effects of Missing Hydrogen Atoms on PDB Based Scatterplots 36 e The fact that hydrogen atoms are
101. uis Missouri USA and are summarised below H hydrogen C 3 C 2 C1 sp sp sp hybridised carbon respectively C ar aromatic carbon C cat cationic e g guanidinium carbon N 3 N 2 sp pyramidal and sp hybridised nitrogen respectively N 4 tetrahedral quaternary nitrogen N ar aromatic nitrogen N am amide nitrogen N pl3 planar trigonal nitrogen 0 3 0 2 sp sp hybridised oxygen respectively O co2 carboxylate phosphate oxygen F Cl Br I fluorine chlorine bromine iodine P 3 phosphorus 5 3 S 2 sp and sp hybridised sulphur respectively S o S 02 sulphoxide and sulphone sulphur respectively Du dummy atom used e g for variable atom types such as the ConQuest X any atom type e Ifthe atom types assigned to central group atoms are incorrect it may affect the identification of topologically equivalent atoms e g the two oxygen atoms of the carboxylate ion and hence the determination of 3D symmetry e By default atom types are derived by IsoGen from the ConQuest output usually with sufficient accuracy to avoid problems in detection of topological and 3D symmetry When problems do occur the user can edit atom types by selecting Atom types in the Expert menu e This creates a pop up window a typical example of which is TsoStar User Guide OK Cancel La l e Clicking on an atom then pops up a small entry box that enables the user to
102. w 1 New Fe fel x File Edit Atoms Bonds 3D Options Help Click and PE ER E to create a bond Next Atom N 2 3D Parameters as cal PE ER E to an existing atom to make a connection Next Bond Single KAA Options CONTACT Delete Contacts Options Delete RingMaker Search biw 4 Store S ic ey s P F CI An More Grou s Nn Bond rel Es Templates cefajofn 3 E 3 Cancel IsoStar User Guide 103 3 Setting up 3D Constraints e Hit ADD 3D This will generate a Geometric Parameters window e Suppose that in our non bonded search we want to find contacts between either the nitrogen or hydrogen atom of the N H fragment and any atom of the methylcarbamoy fragment except for the point of attachment carbon We can do this by specifying that the contact lies between two centroids rather than specific atoms e Hit each atom of the methylcarbamoyl fragment except for the point of attachment carbon 1 e the carbon to which the CONHMe is bonded and pick Define next to Centroid from the Geometric Parameters window You have now defined a centroid CENTI comprising the desired atoms The ConQuest window should look something like this St 5 x Fie Edit Atoms Bonds 3D Wooo Select atoms to define 3D Parameters Defined Centroid CENT1 03 C2 N4 H10 C5 H6 H8 H9 3D Parameters DRA EDIT lly ERASE 12 Fa ADD 3D CONTACT To construct Parameter
103. ween Ligand and Protein We will start off in the Ligand section e Since we are going to investigate carboxylic acids 1 e a terminal group containing only O C and H atoms start by clicking on O C H only in the Terminal section of Ligand TsoStar User Guide RPRNICIF A Home Ligand Terminal Ligand Acyclic Links Ligand Ring Systems Ligand Solvates Protein Plots Custom Plots Help Ligand Terminal O C H only Home e acetoxy Q Ligand Terminal Sa C H only 0 CH N C H only O C H only N O CH only e acetyl Si containing P containing A S containing CH Halo containing carboxylic acid Acyclic links C H only none e charged carboxylic acid NOCH e uncharged carboxylic acid cis P containing e uncharged carboxylic acid trans S containing Ring systems rane Phenyls Q C H only l N C H only al O C H only N O C H only S containing hydroxy Nucleic acid ae bases i e charged aromatic hydroxy e uncharged aliphatic hydroxy Solvates etc Inorganic Organic e uncharged aromatic hydroxy e Scroll through the list of functional groups that appears table These are all the terminal O C H groups for which IsoStar contains information We will start off by looking at carboxylate anions so scroll back towards the top of the list and click on the charged carboxylic acid link e This completes the selection of the central group TsoStar User Guide 93 10 Displa
104. x fold axis along a direction passing through the molecular centroid and perpendicular to mirror plane 1 Note that in axial terms mirror 1 can be regarded as lying along the same axis as the six fold and so one can describe this axis as having 6 m symmetry e Mirror planes perpendicular to the centroid carbon atom vectors passing through the centroid e Assuming the atoms of the benzene ring are numbered to 6 the full point group symmetry can be specified as F Set Symmetry Gm c4ix c3 mi 2 c1 Ok Yew Search Cancel LE e Note here that the c is used to specify the centroid of the benzene fragment 5 1 3 Defining Symmetry Example 3 Sulfate e A sulfate anion has 43m symmetry The 4 axes lie in the directions of the bisectors of the O S O angles the three fold axes lie in the direction of the S O bond vectors the mirror plane axes are perpendicular to the O S O planes e To specify such symmetry three elements are required a 4 axis a three fold axis and a mirror plane For example and assuming the sulfur is atom number and the oxygens atom numbers 2 to 5 SetSymmetry 4 12443 3 12 mea hend Ok Yew Search Cancel CS TsoStar User Guide 65 e The symbol indicates a vector sum in this case the sum of 2 and 3 5 2 Atom types 66 e Each central group atom is assigned an atom type The atom types used are taken from the program SYBYL Tripos Associates St Lo
105. y IMPT method as programmed in TsoStar User Guide 4 6 4 42 the CADPAC 6 0 package This method is free of basis set superposition error a major problem within ab initio supermolecule calculations It provides an estimate for each significant contribution to the total interaction energy viz e The electrostatic energy E term describes the attractive or repulsive classical Coulombic interaction e The exchange repulsion term E 1s the sum of 1 an energy lowering due to exchange of electrons of parallel spin between the molecules and 11 a repulsive term arising from the Pauli exclusion principle e The polarisation energy E term accounts for the energy gain caused by the change of the intramolecular wave function of one molecule due to the presence of the undistorted charge distribution of the second molecule e The charge transfer energy E term is the attractive energy due to actual charge transfer between molecules e The dispersion energy Eg 1s calculated at the second order double excitation level and is the result of instantaneous correlation of fluctuating electron density distributions Due to the limited quality of the empirical repulsion dispersion parameters used in the ORIENT program it is necessary to optimise the energy as a function of the distance between the model molecules maintaining fixed angular orientations using the IMPT method The energies of the resulting distance optimised dimer
106. ying CSD Scatterplots 94 charged carboxylic uncharged carboxylic acid uncharged carboxylic acid acid cis trans charged carboxylic acid PDB scatterplots for all the above groups are identical because the groups are indistinguishable in protein crystal structures More Information General C H only N H O H Other N or O Sulfur Halo halide Amino acid General Links to statistical data Links to theoretical energy data Contact Group CSD PDB Stats Theory any C N O S or H 4923 4996 any polar X H X N O or 2498 4881 This search has not been done General C H only N H O H Other N or O Sulfur Halo halide Amino acid C H only Links to statistical data d Links to theoretical energy data Contact Group CSD PDB Stats Theory any alkyl C H 2498 4897 methylene 965 1643 methyl 952 981 e The new page lists the available contact groups which are displayed in contact group tables The page is subdivided into a number of sections headed General C H only etc The contact group table lists the types of contacts for which IsoStar contains information Launch the CSD based scatterplot for the any C N O S or H contact group 1 e click on the hyperlink in the row marked any C N O S or H and the column headed CSD 4941 in the example above If IsoStar is installed correctly this will launch the IsoStar visualiser displaying a scatterpl

IsoStar User Guide and Tutorials 2010 Release

Contents

Download Pdf Manuals

Related Search

Related Contents