| Jul 1 2004 |
[ vlsoverview | ligdockht | vlsht ] by Max Totrov and Ruben Abagyan
!_ 1. Where to dock. Building Receptor and Pocket Model The goal here is to have an adequate three-dimensional model of the receptor pocket you are planning to dock ligands to. And the pitfalls are that your model is not accurate overall, or does not reflect the induced fit, or alternative conformations of the receptor binding pocket are missed. Receptor from PDB If you have only a single entry with your receptor, convert the protein with convertObject yes yes no no , after deleting water molecules and irrelevant chains (e.g. delete a_!1 ), or use menus as in the ligand docking section. However, if you have a choice between several templates, take the following into account:
Receptor from homology modeling A model by homology can be built with the build model command (menu Homology/Build_Model) followed macro refineModel . Identifying pockets If a binding pocket is not known in advance, use icmPocketFinder or icmCavityFinder (for closed pockets) macros. icmPocketFinder can also be accessed from menu Docking/Receptor Setup , submenu Identify_Binding_Sites !_ 2. What to dock. Ligand, ligands, a database or a library. Usually a good start is to try to dock the known ligand(s) to the receptor model. You may also want to dock a library of compounds in order to identify lead candidates. In this case the main pitfall is that the library is too restricted, molecules are not chemically feasible or not drug-like. Ligand from PDB Then to dock a ligand from pdb, go through the procedure described in the ligand docking section. Ligand(s) from a mol/mol2- file, or SMILES strings. The main prerequisite is that the formal charges and the bond types are correct. If they are not correct, you need to process each molecule manually as described in the ligand docking section. From a command line you may use the build smiles or convert2Dto3D macro. !_ Flexible docking considerations. After the receptor maps are built, you will start a docking simulation. The goal of the flexible docking calculation is prediction of correct binding geometry for each binder. ICM stochastic global optimization algorithm attempts to find the global minimum of the energy function which includes five grid potentials describing interaction of the flexible ligand with the receptor and internal conformational energy of the ligand. During this process a stack of alternative low energy conformations is saved ( one of the choices in the Docking menu ). Some facts about ICM docking:
Pitfalls. Inaccurate receptor model, or incorrectly converted ligands, or insufficient optimization effort may lead to incorrect predictions. !_ Scoring The goal of scoring in virtual ligand screening is to ensure maximal separation between binders and non-binders , and not to rank a small number of binders according to their binding energies. The vls module allows you to access a good scoring function.
ICM ligand docking procedure performs docking of the fully flexible small-molecule ligand to a known receptor 3D structure. Before setting up the docking project, ICM object of the receptor has to be created. In most cases, x-ray structure of the receptor is initially in the PDB format. Thus, it has to be converted to the ICM format. This process involves addition of the hydrogen atoms, assignment of atom types and charges from the residue templates (icm.res) and imposition of internal coordinates tree (icm-tree) on the original pdb coordinates. The easiest way to convert pdb structure into icm object is through GUI as follows:
It is recommended that "optimize hydrogens" option is selected. To accelerate the procedure, disable the 3D graphics window (side menu Clear/No Graphics ) When the procedure finishes, converted object is the 'current' object in icm. You can check the results by displaying the converted structure. !_ Docking project setup Start project setup by defining the project name (menu Docking/Set project name ). Avoid spaces and leading digits in the name. All files related to the docking project will be stored under names, which start from the project name. Most customized parameters will be saved in the table file under the project name as well: DOCK1.tab # control table DOCK1_gb.map # 3D potential grids, or 'maps' DOCK1_gc.map DOCK1_ge.map DOCK1_gh.map DOCK1_gs.map DOCK1_rec.ob # receptor objectetc.. The next step is to set up the receptor (menu Docking/Receptor setup ). Select the receptor molecules, in most cases a_* will do - all molecules in the current object will be included. Define binding site residues, either manually e.g. a_/123,144,152 for selection by residue numbers, or graphically using lasso tool (don't forget to set selection level to residue). This selection is used solely to define boundaries of the docking search and the size of the grids and doesn't have to be complete, selecting some 4 residues delimiting the binding site is sufficient. Receptor setup dialog also lets you run binding site identification routine to quickly locate putative binding sites on your receptor. After the receptor setup is complete, the program normally displays the receptor with the selected binding site residues highlighted in yellow xstick representation. Ligand setup offers a number of ways (submenu docking/ligand setup ) to define the ligand, depending on the source of the ligand structure(s). !_ Converting a chemical from pdb. The Protein data bank, due to unprecedented ignorance, for the last 15 years has not been storing any information about covalent bond types and formal charges of the chemical compounds interacting with proteins! This oversight makes it impossible to automatically convert those molecules to anything sensible and requires your manual interactive assignment of bond types and formal charges for each compound in a pdb-entry. Therefore, if you apply the convert command to a pdb-entry with ligands, the ligands will just become some crippled incomplete molecules which can not be further conformationally optimized. Follow these steps to convert a chemical properly from a pdb form to an a correct icm object.
Setting up a ligand or a set of ligands Let's now consider the situation when icm object of the ligand loaded. ICM object of the ligand can also be prepared, for instance, by reading structure from SD file (menu File/Read Molecule/Mol/SDF ) and converting it to ICM (menu MolMechanics/ICM-Convert/Chemical ). Once the icm object of the ligand is ready, proceed to docking ligand setup (menu Docking/Ligand Setup/From Loaded ICM object ). The ligand setup procedure will first display the grid box, allowing you to adjust the box dimensions, and then the 'probe' which defines the initial positioning of the ligand's center of mass and long/short axis. The probe can be moved/rotated. While its positioning has only minor influence on the results as long as it remains inside the binding site, it may help the procedure to find the correct docked orientation more reliably and/or in shorter time. Ligand setup procedure can be ran repeatedly to change the ligand within the same docking project. Also box size and probe position can be changed later (menu Review/Adjust ligand/box ). At this point, the project is ready for the calculation of maps (menu docking/Make receptor maps ). The calculations generally take several minutes to prepare the maps. While the dialog allows changing the grid step, we do not recommend altering the default value of 0.5 which was found optimal for a large number of test cases. With the map calculations completed, everything is ready to start the actual docking simulation. A larger set of ligands in a mol file can be considered as a database and indexed with the ICM indexing tool (menu Docking/Tools/Index Mol,Mol2 Database ) for fast access. Ligand structures from mol/mol2 file can be converted to ICM on the fly and do not require manual preparations necessary in the case of PDB structures. !_ Running docking simulations Use menu docking/Small Set Docking Batch to start docking of one or few ligands in the background. You can also view the process interactively (menu docking/Interactive Docking ) although it is much slower due to the time spent on drawing the molecules. The results of the batch docking job are saved in the PROJECTNAME_answers*.ob #icm-object file with best solutions for each ligand PROJECTNAME_*.cnf # icm conformational stack files with multiple docked conf. PROJECTNAME_*.ou # output file were various messages are stored.Multiple conformations accumulated during the docking of the ligand can be visualized and browsed in ICM (menu Docking/Browse Stack Conformations ). Use menu Docking/Display/Preferences to change default graphic representation of ligand/receptor. !_ Rundock UNIX shell script Docking batch jobs should be started from the UNIX command line using rundock shell script. The syntax is as follows:
rundock <options> <project name>
options: -f <ligand entry from>" # if using multiple ligand input file
-t <ligand entry to>" # range of ligands to dock can be selected
-L <list of ligands> # comma-separated list of ligand numbers from database
-l <thoroughness> # change the length of MC docking, default is 1.
-n <scanName> # change the run name in the output files
-a # force docking and saving of all compounds
-s # save stack conformations
-S # evaluate score for all stack conformations (slow)
-o # redirect output to <project name>_from-to.ou
-c <output file> # continue interrupted job with <output file>
Example:
rundock -f 3 -t 5 -l 3. -s MYPROJECTwill thoroughly (3 times longer simulation than default) dock ligands 3 to 5 and save conformational stacks for each one of them. !_ Template constrained docking It is possible to use a template object to tether part of the ligand structure to a preferred position during the docking simulation. Prepare an object file containing the group of atoms to be tethered to. Edit the .tab docking setup file, setting the s_templateObj field to the name of the template object file (it is 'none' by default). The variable l_superByName controls the way correspondence is established between the ligand and template atoms. If it is 'no', chemical substructure search is performed and tethers imposed according to the substructure match. If l_superByName is 'yes', simple matching according to atom names is performed. Tethers can be individually weighted by assigning b-factor values to the template atoms. Weights are reversely proportional to b-factor, default b-factor of 20. corresponds to the weight of 1.
#>r DOCK1.r_ScoreThreshold -35. The choice of the threshold can be done in two ways:
!_ Potential of mean force score Potential of mean force calculation ( pmf ) provides an independent score of the strength of ligand-receptor interaction. The pmf-parameters are stored in the icm.pmf file. To enable calculation of the pmf-score, define the PROJECTNAME.r_mfScoreThreshold threshold paramter to the table: #>r PROJECTNAME.r_mfScoreThreshold 999.!_ Other selection criteria ICM VLS uses a number of criteria to pre-select compounds before docking. Edit the project .tab file to change their defaults: #>i DOCK1.i_maxHdonors 5 #>i DOCK1.i_maxLigSize 500 #>i DOCK1.i_maxNO 10 #>i DOCK1.i_maxTorsion 10 #>i DOCK1.i_minLigSize 100!_ Parallelization If the database size exceeds several thousand compounds, it is desirable to run a number of VLS jobs in parallel to speed up calculations. Use -f and -t options of rundock to start multiple jobs on different parts of the database, e.g. rundock -f 1 -t 10000 -o rundock -f 10001 -t 20000 -o rundock -f 20001 -t 30000 -o .. !_ Running VLS jobs in PBS UNIX cluster environment Jobs on the Linux cluster are run through PBS queuing system. Several scripts are provided to facilitate submission of vls jobs. To submit a single job, use pbs script 'pbsrun', which is a pbs wrapper for rundock qsub $ICMHOME/pbsrun -v"JOBARGS=-f 1 -t 1000 -o MYPROJECT" Note that the rundock arguments go in the quotes after JOBARGS= . The qsub command is a part of PBS. To submit multiple jobs, there is a simple shell script 'pbsscan' which executes multiple qsub's for database stripes: $ICMHOME/pbsscan MYPROJECT 1 6000 1000 -submits 6 jobs, 1 to 1000; 1001 to 2000 ... 5001 to 6000. Currently this script only supports default rundock arguments, copy/edit to change. The command qstat is a part of PBS and can be used to check the status of the jobs. In addition, $ICMHOME/scanstat script can be used to monitor the progress of the VLS jobs. It analyses the *.ou rundock output files. $ICMHOME/scanstat *.ou To delete the jobs, use PBS command qdel: qdel 1234 # delets job number 1234 !_ Where to find the scores of the compounds Once the compounds are docked, if VLS option is installed, the procedure evaluates the score and stores it in the 'comment' of the ligand object. When browsing scan answers, the SCORE>... line appears for each object viewed, containing the value of the score and it's component terms. It can also be extracted from the icm object in shell using Namex( a_1. ) function, and Field() can be used to get particular component or the total: Field( Namex( a_1. ) "Score=" 1). The SCORE lines also appear in the output file and can be extracted by simple unix grep command grep SCORE *.ou The MFScore was recently added, it's calculated if r_mfScoreThreshold variable is defined in the project .tab file. It can be added manually: #>r PROJECTNAME.r_mfScoreThreshold 999. !_ Analysis of the results The hits found by the screening procedure and stored in *answers*.ob files can be visualized in ICM (menu Docking/Browse Scan Solutions ). If necessary, they can also be exported as SD file using (menu Docking/Tools/Export scan answers as mol ). The score and its components are stored in the resulting SD file as well. Simple analysis of the score distribution can be performed by making a histogram (menu Docking/Tools/Scan results histogram ).
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