2012 DOCK Tutorial With Streptavidin

2012 DOCK tutorial with Streptavidin - Rizzo Lab
http://ringo.ams.sunysb.edu/index.php/2012_DOCK_tutorial_...
2012 DOCK tutorial with Streptavidin

From Rizzo Lab For additional Rizzo Lab tutorials see DOCK Tutorials. Use this link Wiki Markup (http://www.unilang.org/wiki/index.php/Wiki_markup) as a reference for editing the wiki.
Contents
1 I. Introduction 1.1 DOCK 1.2 Streptavidin & Biotin 1.3 Organizing Directories 2 II. Preparing the Receptor and Ligand 2.1 Downloading the PDB Structure (1DF8) 2.2 Preparing the Ligand and Enzyme in Chimera 3 III. Generating Receptor Surface and Spheres 3.1 Receptor Surface 3.2 Spheres 4 IV. Generating Box and Grid 4.1 Box 4.2 Grid 5 V. Docking a Single Molecule for Pose Reproduction 5.1 Docking and Results 5.2 Results 6 VI. Virtual Screening 6.1 Virtual Screening Preparation 6.2 Virtual Screening Protocol 6.3 Virtual Screening Results 7 VII. Running DOCK in Serial and in Parallel on Seawulf 7.1 Running DOCK in Serial on a Single Processor 7.2 Running DOCK in Parallel using MPI 7.3 Serial Calculation for Pose Reproduction 7.4 Parallel Virtual Screen 8 VIII. Frequently Encountered Problems 8.1 Barbara 8.2 Woo Suk 8.2.1 Distinguishing overlapped spheres 8.3 Longfei 8.4 Roman 8.5 Quan 8.6 Hui 8.7 Tuoling 8.8 Yuchen 8.9 Always Remember Your Path 8.10 Yunting 8.10.1 Delete Spheres Manually 8.11 Kip 8.11.1 Keeping your data in sync 8.11.2 Deleting jobs
I. Introduction
DOCK
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DOCK is a molecular docking program used in drug discovery. It was developed by Irwin D. Kuntz, Jr. and colleagues at UCSF (see UCSF DOCK (http://dock.compbio.ucsf.edu/) ). This program, given a protein binding site and a small molecule, tries to predict the correct binding mode of the small molecule in the binding site, and the associated binding energy. Small molecules with highly favorable binding energies could be new drug leads. This makes DOCK a valuable drug discovery tool. DOCK is typically used to screen massive libraries of millions of compounds against a protein to isolate potential drug leads. These leads are then further studied, and could eventually result in a new, marketable drug. DOCK works well as a screening procedure for generating leads, but is not currently as useful for optimization of those leads. DOCK 6 uses an incremental construction algorithm called anchor and grow. It is described by a three-step process: 1. Rigid portion of ligand (anchor) is docked by geometric methods. 2. Non-rigid segments added in layers; energy minimized. 3. The resulting congurations are 'pruned' and energy re-minimized, yielding the docked congurations.
Streptavidin & Biotin

Streptavidin is a tetrameric prokaryote protein that binds the co-enzyme biotin with an extremely high afnity. The streptavidin monomer is composed of eight antiparallel beta-strands which folds to give a beta barrel tertiary structure. A biotin binding-site is located at one end of each !-barrel, which has a high afnity as well as a high avidity for biotin. Four identical streptavidin monomers associate to give streptavidins tetrameric quaternary structure. The biotin binding-site in each barrel consists of residues from the interior of the barrel, together with a conserved Trp120 from neighboring subunit. In this way, each subunit contributes to the binding site on the neighboring subunit, and so the tetramer can also be considered a dimer of functional dimers. Biotin is a water soluble B-vitamin complex which is composed of an ureido (tetrahydroimidizalone) ring fused with a tetrahydrothiophene ring. It is a co-enzyme that is required in the metabolism of fatty acids and leucine. It is also involved in gluconeogenesis.
Organizing Directories
While performing docking, it is convenient to adopt a standard directory structure / naming scheme, so that les are easy to nd / identify. For this tutorial, we will use something similar to the following:
~username/AMS536/DOCK-Tutorial/00-original-files/ /01-dockprep/ /02-surface-spheres/ /03-box-grid/ /04-dock/ /05-virtual-screen/
In addition, most of the important les that are derived from the original crystal structure will be given a prex that is the same as the PDB code, '1DF8'. The following sections in this tutorial will adhere to this directory structure / naming scheme.
II. Preparing the Receptor and Ligand

Downloading the PDB Structure (1DF8)
The Protein Data Bank contains atomic coordinates for more than 79,000 molecules and is accessible via the internet from PDB (http://www.pdb.org) . The target (protein or nucleic acid) is accessible by PDB ID, molecular name, or author. After entering the PDB ID, molecule name, or author, click Download Files, then select PDB File (text). A window will open; click on Save File Downloads.
Preparing the Ligand and Enzyme in Chimera

If you are on "herbie" you can access Chimera directly by typing in Chimera at the prompt. Otherwise, download Chimera to your desktop and obtain your protein/nucleic acid complex of interest using Fetch (by ID).
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Open Your Protein of Interest and Delete Unwanted Molecules: Click on Open under the File menu and nd where you saved your pdb le. You only need one of the monomers to perform docking. Remove chain B by carrying out the following: 1. Select ! Chain ! B ! All. 2. Actions !Atoms ! Delete. To delete water molecules and/or other ligands, go to Tools ! Structure Edit ! Dock Prep. Check all boxes and click okay. This will walk you through the steps needed to prepare the complex for docking, and will also assign partial charges to the protein and the ligand. Choose am 1-bcc for charges. Save the le as 1DF8.dockprep.mol2. This le contains a conformation of the complex with hydrogen atoms. The grid calculation is based on the receptor with its hydrogen atoms. The grid score is an energy calculation that is based on the following equation: E GRID = E VDW + E ES. The score is an approximation of the molecular mechanics' energy function and it considers only through space interactions. Create a Receptor File: Go to Select ! Residue ! BTN. Then go to Actions ! Atoms ! Delete. Save the le as 1DF8.receptor.mol2. Create a Receptor File with No Hydrogen Atoms: Go to Select ! Chemistry ! Element ! H ! Delete. Save a PDB le as 1DF8.receptor.noH.pdb. Create a Ligand File: Open only the 1DF8.dockprep.mol2. Go to Select ! Chain ! Delete. This will allow you to have only the ligand. Save the le as 1DF8.ligand.mol2. These are the les that you will need to continue with rigid or exible docking.
dockprep image
III. Generating Receptor Surface and Spheres

Receptor Surface
To generate an enzyme surface, rst open the receptor pdb le with the hydrogen atoms removed (1DF8.receptor.noH.pdb). Next, go to Actions -> Surface -> Show. Note that for DOCK calculation hydrogen atoms are considered, but for generating enzyme surface and spheres, it is necessary to use the protein without hydrogens.
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2012_DOCK_Tutorial_1DF8_surface
Recent versions of Chimera include a Write DMS tool that facilitates calculation of the molecular surface. Go to Tools -> Structure Editing -> Write DMS. Save the surface as 1DF8.receptor.dms.
2012_DOCK_Tutorial_1DF8_dms
The Write DMS tool will "roll" a small probe (default radius = 1.4 Angstroms = Size of a water molecule) over the surface of the enzyme and calculate the surface normal at each point. DMS (distributed molecular surface) le is subsequently used as input le for sphgen.
Spheres
To generate docking spheres, we need to use a command line program called sphgen. To run the sphgen, we need a input le named INSPH.
1DF8.receptor.dms R X 0.0 4.0 1.4 1DF8.receptor.sph #molecular surface file that we got from previous step #sphere outside of surface (R) or inside surface (L) #specifies subset of surface points to be used (X=all points) #prevents generation of large spheres with close surface contacts(defalut=0.0) #maximum sphere radius in angstroms (default=4.0) #minimum sphere radius in angstroms (default=radius of probe) #clustered spheres file that we want to generate
Save the INSPH le. Then use this command to generate the spheres le:
sphgen -i INSPH -o OUTSPH
-i means input; -o means output. You should get two output les: OUTSPH and 1DF8.receptor.sph. The OUTSPH le should similar to this:
density type = X reading 1DF8.receptor.dms # of atoms = 881 # of surf pts = 10771 finding spheres for 1DF8.receptor.dms dotlim = 0.000
type
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radmax = 4.000 Minimum radius of acceptable spheres? 1.4000000 output to 1DF8.receptor.sph clustering is complete 28 clusters
You can also open the spheres le that we generated in this step (1DF8_receptor.sph). This le contains detailed information about the spheres, which are divided into 28 clusters. Cluster 0 in the end of the spheres le is a combination of all the clusters. In order to visualize the generated spheres, you can use a program called showsphere. Showsphere is an interactive program. In the command line, simply type
showsphere
You will be asked the following questions:

Enter name of sphere cluster file: 1DF8.receptor.sph Enter cluster number to process (<0 = all): -1 Generate surfaces as well as pdb files (<N>/Y)? N Enter name for output file prefix: 1DF8.output. Process cluster 0 (contains ALL spheres) (<N>/Y)? N
In this case, 28 pdb les should be generated. An alternative way to do this is to create an input le, showsphere.in, as follows:
1DF8.receptor.sph -1 N 1DF8.output N
Then type the command showsphere

showshpere < showsphere.in
After generating the pdb les by showsphere, you can visualize each cluster by UCSF Chimera. For example, open 1DF8.output_1.pdb by Chimera, then choose Actions->Atoms/Bonds->sphere, you will be able to see the spheres in cluster 1. You can also open the receptor le (1DF8.receptor.mol2) at the same time, then choose Presets->Interactive 3(hydrophobicity surface), then again choose Actions->Atoms/Bonds->sphere, you will be able to see what the spheres in cluster 1 look like on the enzyme surface.
2012_DOCK_Tutorial_1DF8_spheres
There are over 500 spheres in the spheres le(1DF8.receptor.sph). However, we're only interested in docking the ligand into the active site. Therefore we need to select only those spheres which are inside the active site, using sphere_selector program.
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sphere_selector 1DF8.receptor.sph ../01-dockprep/1DF8.ligand.mol2 10.0
Sphere_selector lters the output from sphgen, selecting all spheres within a user-dened radius of a target molecule. In this case, we selected the spheres within 10 angstroms of our ligand. A le called selected_spheres.sph should be created, showing the spheres that are selected. You can again visualize it by showsphere. You can also change the radius (to 8 angstroms or 6 angstroms) or manually edit the le selected_spheres.sph so that you can select the spheres you want. In this tutorial, spheres within 6 angstroms of the ligand are used for the next step (with one sphere which does not belong to the active site being manually deleted).
2012_DOCK_Tutorial_1DF8_surface_spheres_10 angstroms
2012_DOCK_Tutorial_1DF8_surface_spheres_6 angstroms
IV. Generating Box and Grid

Box
In order to speed up docking calculations, DOCK generates a ne grid. At each point in the grid, electrostatic and the VDW probes' energies are precomputed. The energies are computed using a molecular force eld. To determine the dimensions of the grid, however, we rst generate a box that contains the outer boundaries for the grid calculation. The dimensions and location of the box can be determined using a program called showbox. First create a directory where you will place the grid les.
$mkdir 03-box-grid $cd 03-box-grid
showbox can be used interactively or a le with predetermined answers can be fed into the program. The program asks the questions depicted in the diagram the right: To run the program in the interactive mode, run
$showbox
To feed the answers to the questions, run

$showbox < showbox.in
For example, showbox.in can contain:

Y 5 ../02-surface-spheres/selected_spheres.sph 1 1DF8.box.pdb
Flow Chart of Questions for Showbox (Red path is followed
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in this tutorial) Y means we use automatic box construction, 5 is the extra margin to be enclosed around our ligand (in Angstroms), selected_spheres.sph is the sphere le we generated, 1 corresponds to the cluster number in the selected_spheres.sph le, and 1DF8.box.pdb is the output le. We can open the output box le in chimera to make sure the box is in the right place.
Grid
Now let's generate a grid within our box. We will use the energy scoring method to generate a grid, resulting in three additional les with extensions *.nrg, *.bmp, and *.out. The *.nrg le contains the energy scoring, *.bmp contains the size, position and grid spacing and determines whether there are any overlaps with receptor atoms. To generate the grid we will use the grid program. This program can either be used interactively, or an input le can be fed in, just like the showbox program.
Usage: grid [-i [input_file]] [-o [output_file]] ... [-standard_i/o] [-terse] [-verbose] -i: read from grid.in or input_file, standard_in otherwise -o: write to grid.out or output_file (-i required), standard_out otherwise -s: read from and write to standard streams (-i and/or -o illegal) -t: terse program output -v: verbose program output
1DF8 receptor along with our ligand and the box we generated using showbox
For our grid.in le, we will use the following answers:

compute_grids grid_spacing output_molecule contact_score energy_score energy_cutoff_distance atom_model attractive_exponent repulsive_exponent distance_dielectric dielectric_factor bump_filter bump_overlap receptor_file box_file vdw_definition_file score_grid_prefix yes 0.3 no no yes 9999 a 6 9 yes 4 yes 0.75 ../01-dockprep/1DF8.receptor.mol2 1DF8.box.pdb /opt/software/AMS536software/dock6/parameters/vdw_AMBER_parm99.defn grid
Line by line: 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. compute scoring grids (yes) what is the distance between grid points along each axis (in Angstroms). write up coordinates of the receptor into a new le compute contact grid? default is no compute energy score? yes - we are using this method to compute force elds on probes the max distance between atoms for the energy contribution to be computed atom_model u means united atom model where atoms are attached to hydrogens, and a stands for all-atom model, where hydrogens on carbons are treated separately attractive component stands for exponent of the attractive LJ term in VDW potential repulsive component stands for exponent in the repulsive LJ term in VDW potential distance dielectric stands for the dielectric constant to be linearly dependent on distance distance dielectric factor is the coefcient of the dielectric bump lter ag determines if we want to screen orientation for clashes before scoring and minimization bump_overlap stands for the fraction of allowed overlap where 1 corresponds to no allowed overlap and 0 corresponds to full overlap being permitted. our receptor le the box le we generated in the Box section
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16. VDW parameters le 17. Prex for the grid le name. All the extensions will be generated automatically.
V. Docking a Single Molecule for Pose Reproduction

Docking and Results
Change directory into 04-dock and create an empty input le called dock.in
$touch dock.in #create a file called dock.in
Run dock6.4
$dock6 -i dock.in
Alternatively:
$dock6 -i dock -v # -v option allows you to print out the information regarding each growth step on terminal
Or:
$dock6 -i dock -o dock.bf.out -v # -o allows you to write the aforementioned information into an output file named dock.bf.out
1st run: Notice that running dock6 with an empty input le will require you to answer a series of questions. For the rst run we will deactivate most of the features by selecting no. Parameters requiring specication are listed below:
ligand_atom_file [database.mol2] (): ligand_outfile_prefix [output] (): ../01-dockprep/1DF8.ligand.mol2 1DF8.output
What the program is doing in the 1st run is to take the ligand.mol2 le and directly generate an output le without any additional manipulations. You would expect it to be exactly the same as the original pose. You can open it in Chimera by using the ViewDock function under Tools->Surface/Binding Analysis. We will not show the result here. 2nd run: The real experiment begins here. Notice that we selected no for most of the functions. This time we will try change some parameters in the dock.in le.
$vim dock.in
Parameters being changed are listed below:

orient_ligand yes
The "orient_ligand" option tells the program whether to try orientations different from the pose in the original .mol2 le. Note that because of the change, additional questions are asked by the program. For simplicity we keep most of the answers as default. File paths that need to be specied are listed below:
receptor_site_file vdw_defn_file flex_defn_file flex_drive_file [receptor.sph] ():../02-surface-spheres/selected_spheres_06A.sph [vdw.defn] ():../03-grid/vdw_AMBER_parm99.defn [flex.defn] ():../03-grid/flex.defn [flex_drive.tbl] ():../03-grid/flex_drive.tbl
The receptor_site_le should be the selected spheres le (.sph) generated in a previous step (02 surface and spheres), according to which the program orients the ligand.
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The vdw_defn_le instructs the dock6 program to use the Van der Waals parameter sets from the AMBER force eld. The ex_defn_le and the ex_drive_le contain the information required by the program to sample conformations. The result is shown below. As you may notice the result doesn't look very good.
DOCK second Run result
3rd Run: Here we will further specify parameters to improve the result.
calculate_rmsd score_molecules num_scored_conformers yes yes 10
Again, further questions will be asked when you run the program. Keep most answers as the default values. Paths requiring specication are listed below:
grid_score_grid_prefix [grid] ():../03-grid/1DF8.grid
Note that the above specication will tell the program to load the 1DF8.grid.nrg le you generated in the previous step (grid) for scoring.
simplex_max_iterations write_conformations cluster_rmsd_threshold [1000] ():20 [yes] (yes no):no [2.0] ():0.2
The simplex_max_iteration parameter species number of minimization cycles. Note the program will cluster poses that are very close together (rmsd smaller than the threshold specied in the cluster_rmsd_threshold parameter) into a cluster. This time the program returned the best 10 poses. The one with the best grid score (-52.8, shown blue) superimposed quite well with the original pose in the crystal structure (RMSD = 0.71). You can view the grid scores in ViewDock by selecting Column->Show->Grid Score
DOCK third Run result
4th Run: This time we will set the ligand as exible:
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flexible_ligand
yes
Further questions will be asked during the run. Keep most of the answers as the default values except for:
simplex_grow_max_iterations [20] ():500
The simplex_grow_max_iteration species the maximum number of iterations per cycle per growth step. Notice that the run is signicantly slower this time. Again the best pose generated (-64.8) is shown in blue. The grid score improved signicantly but the RMSD did not change much (0.79)
DOCK fourth Run result
5th Run: This time we will turn the bump lter on:
bump_filter yes
The bump_lter option lters out conformations that cause clashes between atoms. Further questions will be asked during the run. Keep answers as the default values. Specify the following path:
bump_grid_prefix [grid] ():../03-grid/1DF8.grid
Note that the path tells the program to access the .bmp le generated in the previous grid step. The best pose (grid score -65.3) is again shown in blue. Notice that turning on the bump lter does not alter either the grid score or the RMSD (0.78).
DOCK fth Run result
Remember you can tweak any parameter in the dock.in le instead of keeping them as default. Following is the nal dock.in le used during the fth run:
ligand_atom_file limit_max_ligands skip_molecule read_mol_solvation calculate_rmsd use_rmsd_reference_mol ../01-dockprep/1DF8.ligand.mol2 no no no yes no
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use_database_filter orient_ligand automated_matching receptor_site_file max_orientations critical_points chemical_matching use_ligand_spheres use_internal_energy internal_energy_rep_exp flexible_ligand min_anchor_size pruning_use_clustering pruning_max_orients pruning_clustering_cutoff pruning_conformer_score_cutoff use_clash_overlap write_growth_tree bump_filter score_molecules contact_score_primary contact_score_secondary grid_score_primary grid_score_secondary grid_score_rep_rad_scale grid_score_vdw_scale grid_score_es_scale grid_score_grid_prefix dock3.5_score_secondary continuous_score_secondary gbsa_zou_score_secondary gbsa_hawkins_score_secondary amber_score_secondary minimize_ligand minimize_anchor minimize_flexible_growth use_advanced_simplex_parameters simplex_max_cycles simplex_score_converge simplex_cycle_converge simplex_trans_step simplex_rot_step simplex_tors_step simplex_anchor_max_iterations simplex_grow_max_iterations simplex_grow_tors_premin_iterations simplex_random_seed simplex_restraint_min atom_model vdw_defn_file flex_defn_file flex_drive_file ligand_outfile_prefix write_orientations num_scored_conformers write_conformations cluster_conformations cluster_rmsd_threshold rank_ligands
no yes yes ../02-surface-spheres/selected_spheres_06A.sph 500 no no no yes 12 yes 40 yes 100 100 25 no no no yes no no yes no 1 1 1 ../03-grid/1DF8.grid no no no no no yes yes yes no 1 0.1 1.0 1.0 0.1 10.0 500 500 0 0 no all ../03-grid/vdw_AMBER_parm99.defn ../03-grid/flex.defn ../03-grid/flex_drive.tbl 1DF8.output no 10 no yes 0.2 no
Now we will write up a script for submitting your dock job to Seawulf. Create a script called sub.dock.csh
$vim sub.dock.csh
wherein you write:

#! /bin/tcsh #PBS -l nodes=1:ppn=1 #PBS -l walltime=01:00:00 #PBS -o zzz.qsub.out #PBS -j oe cd ~/AMS536/DOCK_tutorial/05-dock_qsub /nfs/user03/sudipto/dock6/bin/dock6 -i dock.in -o dock.out
This will request 1 processor from the cluster for your job. When you are submitting the job:
$qsub sub.dock.csh
Note that you might have to make the script executable before running it:
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$chmod +x sub.dock.csh
Results
VI. Virtual Screening

Virtual Screening Preparation
Virtual screening is a widely used method in computational drug design. It searches large libraries of chemical compounds to identify favorable structures that bind to a target molecule. To perform virtual screening, we use ligands.mol2, a mol2 le which contains 101 small molecules to be the virtual library. The computational cost is reasonable for a quick search. Usually we use larger molecule set from chemical database, such as ZINC(http://zinc.docking.org/). Since the computational cost of virtual screening is much higher than single-ligand docking, it is better for us to run it on Seawulf. We need to compress the whole DOCK-Tutorial directory, copy it to Seawulf and compress it.
tar -cvf DOCK-Tutorial.tar DOCK-Tutorial/ scp DOCK-Tutorial sw:/nfs/user03/usrname/AMS536 tar -xvf DOCK-Tutorial.tar DOCK-Tutorial
Virtual Screening Protocol

The purpose of virtual screening is different from single molecule docking, so we need to modify our previous docking script dock.in to vs.in. We can see the difference between the two les.
1c1 < ligand_atom_file --> ligand_atom_file 56,59c56,59 < vdw_defn_file < flex_defn_file < flex_drive_file < ligand_outfile_prefix --> vdw_defn_file > flex_defn_file > flex_drive_file > ligand_outfile_prefix 61c61 < num_scored_conformers --> num_scored_conformers 63c63,64 < cluster_conformations --> cluster_conformations > cluster_rmsd_threshold
ligands.mol2 ../01-dockprep/1DF8.ligand.mol2 /nfs/user03/sudipto/dock6/parameters/vdw_AMBER_parm99.defn /nfs/user03/sudipto/dock6/parameters/flex.defn /nfs/user03/sudipto/dock6/parameters/flex_drive.tbl 1DF8.vs.output ../03-box-grid/vdw_AMBER_parm99.defn ../03-box-grid/flex.defn ../03-box-grid/flex_drive.tbl 1DF8.output 1 10 no yes 0.2
num_scored_conformers: 1 -> 10 In virtual screening, we only need the most favorable pose of each candidate molecule and compare them. cluster_conformations: yes -> no Slightly different conformations are not clustered together. They are treated as different conformations in the docking process. In order to generate different search spaces, we can modify some other parameters. max_orientations: The maximal number of anchor orientations that will be generated. min_anchor_size: The minimum number of atoms for an anchor. pruning_use_clustering: Pruning conformations during the clustering process. use_internal_energy: Using repulsive VDM to avoid internal clashes. Parallel computing can reduce the running time of virtual screening. Here is our job submission script sub.virtual_screen.csh.
#! /bin/tcsh #PBS -l nodes=4:ppn=2 #PBS -l walltime=01:00:00 #PBS -o zzz.qsub.out
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#PBS -j oe #PBS -V set nprocs = `wc -l $PBS_NODEFILE | awk '{print $1}'` echo "Running on ${nprocs} processors" echo "" echo "processor list are:" cat $PBS_NODEFILE cd ~/AMS536/DOCK_Tutorial/06-virtualscreen mpirun -np $nprocs dock6.mpi -i vs.in -o vs.out
Finally, we can submit the job and perform virtual screening.

qsub sub.virtual_screen.csh
Virtual Screening Results

After performing virtual screening on Seawulf or any other computer, if you are using a parallel computer, you should get a multi-mol2 le 1DF8.vs.output_scored.mol2 which contains the mol2 les of all succesfully docked ligands, a vs.out le which contains the dock results of successfully docked ligands, and vs.out.1 through vs.out.7 which contain dock results from different nodes you use (the number will vary according to the number of nodes you use, here we use 8 nodes as mentioned before). The vs.out le is returned by the leading node and contains the information of each succesfully docked ligands, looks like this:
Molecule: ZINC33171556 Anchors: Orientations: Conformations: Grid Score: Grid_vdw: Grid_es: Int_energy: 1 500 116 -52.373383 -51.643253 -0.730129 6.125062
The vs.out.1 through vs.out.2 les are returned by other nodes processing those ligands, separately. For those succesfully docked ligands, the le will return the elapsed time for docking, and for those not succesfully docked, it will return an error massege like this:
Molecule: ZINC20605433 Elapsed time: ERROR: 0 seconds
Could not complete growth. Confirm that the grid box is large enough to contain the ligand, and try increasing max_orientations.
If you download the multi-mol2 le from seawulf using:

scp sw:~/AMS536/DOCK_tutorial/06-virtual-screen/1DF8.vs.output_scored.mol2 ./
Now you have this multi-mol2 le 1DF8.vs.output_scored.mol2 on your local machine, you can actually open this le in Chimera to visually check you docking results and do some visual analysis. But it's not a good idea to directly open your multi-mol2 le because it contains information of all 47 succesfully docked ligands, if you just open this le, it will be pretty messy. So what you will do is, rst you open the receptor le 1DF8.receptor.mol2 and the ligand 1DF8.ligands.mol2 as a reference. And then use the ViewDock function of Chimera to look at your 47 ligands one or however many you want at a time. You can nd ViewDock via Tools -> Surface/Binding Analysis -> ViewDock, and then nd your 1DF8.vs.output_scored.mol2 le in your own directory and click on open. Now a new window of ViewDock will pop out and there will be an extra ligands in Chimera main window which is the highlighted ligand in ViewDock window. You can look at the ligands one at a time, you can also hold ctrl and click on different ligands to view them at the same time, this will give you a direct idea of how good these ligands dock. The other thing you can do is, the multi-mol2 le 1DF8.vs.output_scored.mol2 contains the energy score of every ligand, so in ViewDock window, you can go to Column -> show -> Grid Score to show the grid score, and then you can click on the head of the column to rank order all the ligands by their grid scores.The picture showing here is the best and worst scored ligands of out calculation, the best scroed one in cyan and the worst scored on in magenta, and the reference ligand is colored according to elements. As you can see, the best scored ligand ts in the binding pocket very well, but the worst scored one almost sticks out of the binding pocket.
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Example of Virtual Screening Result
VII. Running DOCK in Serial and in Parallel on Seawulf

The Seawulf Cluster has 235 dual processor nodes (2 processors per node), for a total of 470 individual processors. These are 3.4Ghz Intel Pentium IV Xeon processors from Dell. Each node has 2GB attached RAM and a 40GB hard disk. Typically you will use one processor on a single node to dock one ligand. If you are docking multiple ligands, you can use more than processor in parallel mode, but you should never use more processors than you have ligands.
Running DOCK in Serial on a Single Processor

The following sample code can be used to run DOCK on one processor on a single node:
#!/bin/tcsh #PBS -l nodes=1:ppn=1 #PBS -l walltime=01:00:00 #PBS -N dock6 #PBS -M user@ic.sunysb.edu #PBS -j oe #PBS -o pbs.out cd /nfs/user03/sudipto/DOCK_Tutorial /nfs/user03/sudipto/dock6/bin/dock6 -i dock.in -o dock.out
Here is an explanation of the code and format:

#!/bin/tcsh #PBS -l nodes=1:ppn=1 #PBS -l walltime=01:00:00 #PBS -N dock6 #PBS -M user@ic.sunysb.edu #PBS -j oe #PBS -o pbs.out cd /path-to-your-home-directory-on-seawulf/DOCK_Tutorial /nfs/user03/sudipto/dock6/bin/dock6 -i dock.in -o dock.out #Execute script with tcsh #Use one node, and one processor per node, so one single processor #Allow 1 hour for your job run #Name of your job #Get an email notifying you when your job is completed #Combine the output and error streams into a single output file #Name of your output file #Change to your home directory and folder with dock files #Specifies path to dock executable and provide input and output filenames
Running DOCK in Parallel using MPI

The following sample code can be used to run DOCK on 4 nodes, using both processors on each node, for a total of 8 processors.
#! /bin/tcsh #PBS -l nodes=4:ppn=2 #PBS -l walltime=24:00:00 #PBS -o zzz.qsub.out #PBS -j oe #PBS -V set nprocs = `wc -l $PBS_NODEFILE | awk '{print $1}'`
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echo "Running on ${nprocs} processors" cd /nfs/user03/sudipto/DOCK_Tutorial cp /nfs/user03/sudipto/dock6/parameters/vdw_AMBER_parm99.defn . cp /nfs/user03/sudipto/dock6/parameters/flex* . mpirun -np $nprocs /nfs/user03/sudipto/dock6/bin/dock6.mpi -i dock.in -o dock.out
For more information, see PBS Queue
Serial Calculation for Pose Reproduction Parallel Virtual Screen
VIII. Frequently Encountered Problems

Barbara
How to Open Files It is important to remember that if you save les to your desktop as mol2 les or pdb les when preparing your ligand and receptor for docking, they cannot be opened using Windows because Windows only opens pdf les using Adobe Reader. You can only access these les through Chimera. If you want to view a le in chimera, rst transfer the le from "herbie" or "seawulf" to your laptop using WinSCP. Then open Chimera on your laptop to open the les.
Woo Suk
Distinguishing overlapped spheres Sometimes your 1DF8.output_1.pdb can have some overlapped spheres. In this case, you cannot nd them before you delete one sphere and you have to repeat this one more time to delete another one. In order to avoid the situation, you can adjust a parameter in INSPH le like following:
1DF8.receptor.dms R X 0.001 4.0 1.4 1DF8.receptor.sph
You can change the fourth parameter from 0.0 to other values such as 0.1, 0.01, or 0.001. Because what you want is just to distinguish the very close spheres, you don't need large numbers.
Longfei
Installation of DOCK If you want to install DOCK on your own desktop or laptop, and you are a beginner of Unix/Linux, you might encounter some problems during installation. A good reference for the installation is the DOCK User Manual(http://dock.compbio.ucsf.edu/DOCK_6/dock6_manual.htm). The problem I encountered was: after I used gnu as conguration le to congure the Makele, I successfully made the cong.h le, but when I was trying to build the DOCK executables, an error message " g77 command not found " appeared. And it is quite inconvenient to install the g77 command. A way to solve this problem is to open the cong.h le, and manually change the g77 in that le to gfortran. Use the Correct Path of Your File One frequently encountered problem is using the wrong path of your le. For example, when you use sphere_selector to generate spheres around the ligand, it requires two les--one is the spheres le .sph and another is your ligand le, but your ligand le is probably not in your working directory! So you need either copy your ligand le to the current directory or specify the correct path of your ligand le like below:
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sphere_selector 1DF8.receptor.sph ../01-dockprep/1DF8.ligand.mol2 10.0 This is also important when you run DOCK. Remember to use the correct path and name of the your le. Notice the Shell That You are Using This tutorial is based on the shell tcsh, and maybe you are using a different shell. This will not cause any trouble in most of the time, but you may want to notice this in some circumstances, for example when you want to execute some .csh le. One way to solve this problem is that simply type "tcsh" in the command line, and you will be running tcsh in another shell.
Roman
One of the problems one may encounter with grid creation is the sampling location for ligand binding. It is important to create a grid and dock the ligand to the correct location in the protein. If the grid is created in the incorrect location, the ligand binding sites will be sampled within the grid, and the best binding site will not be sampled. In our case, we are docking it to an area where a ligand is known to bind to streptavidin. However, this may not always be the case. One solution is to dock to the center of mass of the spheres clusters created with spheregen and assess the center of mass as viable or not based on your chemical intuition. Another way is to assess the protein for buried sites where there may also be a number of spheres. Both are options in the showbox program. Finally, one can attempt to nd proteins with similar sequence and determine where their ligand binding sites are, and then pick the binding site in the protein you are docking to using that information.
Quan Hui
Keep the previous dock6 output .mol2 les When we run dock6 program, the output le would be "1DF8.output_scored.mol2", with the prex "1DF8.output" that you specied in dock.in le. In the case that you would have several runs of dock6 program with the output le of the same path (in the same directory), and you want to keep the result for each run, you need to rename "1DF8.output_scored.mol2" le to another one, otherwise the old .mol2 le will be overwritten by the new .mol2 le generated in the next run. For example, you can rename the old .mol2 le generated by the rst run and then perform the second run.
$mv 1DF8.output_scored.mol2 first_1DF8.output_scored.mol2
Tuoling
Some recommended parameters we can change when running dock
max_orientations min_anchor_size pruning_conformer_score_cutoff pruning_clustering_cutoff 500 40 25 100
Explanations of these parameters can be found on the website provided by Long Fei. Increase the number of these parameters can lead to increased running time. But it enables more thorough cycling and may give you better results.
Yuchen Always Remember Your Path

One situation often appears is that the program will return an error saying "Could not open 1DF8.ligands.mol2 for reading." or something like that indicating that the program couldn't nd the input le you specied in you dock.in or vs.in or some other les. In this case, the most often reason is that the path you specied in your input le is wrong. So the rst thing you might want to do is to check those paths, pay special attention to those paths where you use "." to indicate the same folder
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and ".." to indicate the folder up one level because this is the place where you are most likely to make mistakes. If you are not sure of the usage of "." and "..", it will be safer for you to use the absolute paths of the les. And you can use the command pwd to ask the system where you are if you don't know that.
Yunting
Delete Spheres Manually We need to play some tricks if we want to manually select the spheres in selected_spheres.sph. 1. Transfer sph le to pbd le by using showsphere. 2. Open the pdb le in Chimera and choose the sphere we want to delete. 3. Identify the number of that sphere by Actions->Label->residue->specier. 4. Open sph le, delete that sphere and modify the number of spheres in that cluster.
DOCK spheres within 6.0 ang of ligands cluster 1 number of spheres in cluster 60 28.86000 11.09173 4.97943 2.337 586 67 28.67840 9.74620 12.13940 1.400 60 161 27.11509 13.42709 13.48051 1.401 385 174 27.79940 12.82468 12.04078 2.475 480 183 34.13903 14.01393 7.31591 3.590 691 187 36.78641 17.02434 9.54436 3.481 691 385 30.01753 14.79655 14.46633 1.402 174 463 25.74631 14.83304 9.98248 1.400 589
22 0 0 0 0 0 0 0 0
0 0 0 0 0 0 0 0
Kip
Keeping your data in sync One problem you may encounter is that you want to run your jobs on Seawulf, but you also want the same les on a mathlab computer (or your home computer) for analysis. One option is to use RSYNC to keep your les on Seawulf and Herbie (or any mathlab computer) in sync. For example, if you want to move your DOCK Tutorial les to Seawulf, do the following from Herbie or any mathlab computer:
rsync -arv /home/username/DOCK_Tutorial/ sw:/nfs/user03/username/DOCK_Tutorial
Note: The trailing slash on /home/username/DOCK_Tutorial/ means that rsync will only copy the contents of your DOCK_Tutorial folders. If you want to copy the DOCK_Tutorial folder itself, as well as its contents, then remove the trailing /. To copy newer les from Seawulf back to Herbie, do the following from Seawulf:
rsync -arv /nfs/user03/username/DOCK_Tutorial/ username@compute.mathlab.sunysb.edu:/home/username/DOCK_Tutorial
You can apply this same strategy to then sync les with your home computer as well. Use the following format:
rsync -arv source target
Note: this is easiest if you are using a Linux or Mac computer at home. Deleting jobs If you make a mistake and need to delete a job from the queue, rst list all your queued or running jobs:
qstat -u kip
IMPORTANT: If you get no output from qstat, it means that whatever jobs you have submitted are done! If you do have jobs running or waiting in the queue, qstat will output a list that includes their job id(s). Find the job id for the one you want to delete, and do:
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qdel jobid
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2012 DOCK Tutorial With Streptavidin

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