Difference between revisions of "2018 DOCK tutorial 1 with PDBID 2NNQ"

From Rizzo_Lab
Jump to: navigation, search
(Energy minimization)
(Selecting Spheres)
Line 94: Line 94:
This command will select all of the spheres within 10.0 angstroms of the ligand and output them to selected_spheres.sph. Visualize the selected spheres using Chimera to make sure the correct spheres are selected.
This command will select all of the spheres within 10.0 angstroms of the ligand and output them to selected_spheres.sph. Visualize the selected spheres using Chimera to make sure the correct spheres are selected.
[[File:2nnq selected.png|thumb|center|1000px|2nnq receptor and selected spheres]]
=IV. Generating box and grid=
=IV. Generating box and grid=

Revision as of 16:26, 9 February 2018

This tutorial contains, a step by step by approach to dock a known ligand to a known receptor.

I. Introduction


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). 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 configurations are 'pruned' and energy re-minimized, yielding the docked configurations.


The tutorial will be based on the PDB file 2NNQ downloaded from the PDB Database. 2NNQ is the crystal structure for a human adipocyte fatty acid binding protein in complex with ((2'-(5-ethyl-3,4-diphenyl-1H-pyrazol-1-yl)-3-biphenylyl)oxy)acetic acid.

Organization of Directories

Maintaining a clearly organized set of folders will be helpful in finding specific files, calling different files in input files and most importantly keeping track of everything you do. We would like to recommend to maintain the following set of files throughout the tutorial.


II. Preparation of the ligand and receptor

Download the pdb file 2NNQ from PDB database save it in 0.files folder.

Checking the structure

 - Read the article related to the PDB file to understand protonation states, charges, environmental conditions and other important information regarding the receptor and the ligand.
 - Open the pdb file through chimera and look at the structure. Identify the main components of the model (receptor, ligand, solvent, surfactants, metal ions)
 - Carefully look to identify if there are any missing residues or missing loops. (This particular PDB file didn't contain any missing loops or missing residues)

Preparation of receptor

 - Open the PDB file (2NNQ.pdb) via Chimera
 - Isolate the receptor using select tool and delete tool in Chimera.
 - Save the isolated receptor as a mol2 file. (2nnq_rec_noH.mol2)
 - Open 2nnq_rec_noH.mol2 file again using Chimera and use the following instructions to prepare the receptor file to be used in DOCK.
          Tools -> Structure Editing -> Add H (To add Hydrogen atoms)
          Tools -> Structure Editing -> Add Charge (To add the charge use the latest AMBER force filed available for standard residues. Here we used AMBER ff14SB)
          Save as a mol2 file. (22nq_rec_withH.mol2)
 - If you follow the step below all the above stated steps will automatically appear one after the other to prepare the receptor. 
          Tools -> Structure/Binding Analysis -> DockPrep

Preparation of ligand

 - Open the PDB file via Chimera.
 - Using Chimera, isolate the ligand, add H atoms, add charge and save it as a mol2 file by following the same steps followed for the receptor.

Once all the files are prepared make sure to save the files in 1.dockprep folder.

III. Generating receptor surface and spheres

Preparation of DMS file

 - Open 2nnq_rec_noH.mol2 using chimera.
 - Action -> Surface -> Show
 - Tools -> Structure Editing -> Write DMS
 - Save the 2nnq_rec_withH.dms into 3.surface_spheres folder

Transfer all the folders created so far to seawulf cluster to be used in DOCK.

Generating spheres

 - Go to 2.surface_spheres folder
 - Create a new input file to create spheres by typing vim INSPH and type the following lines inside the file. 

The first line 2nnq_rec_noH.dms specifies the input file. R indicates that spheres generated will be outside of the receptor surface. X specifies all the points will be used. 0.0 is the distance in angstroms and it will avoid steric clashes. 4.0 is the maximum surface radius of the spheres and 1.4 is the minimum radius in angstroms.The last line 2nnq_spheres.sph creates the sph file that contains clustered spheres.

Once the INSPH file is ready, type the following command to generate the spheres.

 sphgen -i INSPH -o OUTSPH

Once sphgen command is successful, 2nnq_spheres.sph file will be created. Open it up using Chimera along with 2nnq_rec_noH.mol2 file. You should get a similar output like the image below.

Selecting Spheres

Here we will be selecting the spheres which defines the binding pocket of the ligand because we are trying to direct the ligand towards that binding site rather than all over the receptor. To select the spheres type the following command.

 sphere_selector 2nnq_rec.sph ../1.dockprep/2nnq_lig_withH.mol2 10.0

This command will select all of the spheres within 10.0 angstroms of the ligand and output them to selected_spheres.sph. Visualize the selected spheres using Chimera to make sure the correct spheres are selected.

2nnq receptor and selected spheres

IV. Generating box and grid

Generating box

Move to 3.boxgrid directory Create a new file showbox.in and write the following lines in the file.


Each of the above lines indicate that;

 We intend to generate a box
 The box length should be 8 Angstroms
 Use the selected_spheres file in the designated location
 The name of the file that contains generated box.

Use the following command to generate the box.

 showbox < showbox.in

If this step is successful, you should see a new file (2nnq.box.pdb) in 3.boxgrid folder.

Generating grid

Create a new file (grid.in)

Use the following command to generate the grid.

 grid -i grid.in -o gridinfo.out

Answer the prompted questions with the answers given below. (or you can use the following lines and include them in the grid.in file before entering the above command. If you do that these questions won't be prompted again. They will be automatically answered by grid.in file created)

compute_grids                             yes
grid_spacing                              0.4
output_molecule                           no
contact_score                             no
energy_score                              yes
energy_cutoff_distance                    9999
atom_model                                a
attractive_exponent                       6
repulsive_exponent                        12
distance_dielectric                       yes
dielectric_factor                         4
bump_filter                               yes
bump_overlap                              0.75
receptor_file                             ../1.dockprep/2nnq_rec_withH.mol2
box_file                                  2nnq.box.pdb
vdw_definition_file                       /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn
score_grid_prefix                         grid

If the command is successful, three new files will be generated. (gridinfo.out, grid.nrg, grid.bmp). Go through gridinfo.out file to make sure all the information about the receptor in the file matches with the original information of the receptor. (Eg:- Total charge, residues and their charges) If the information doesn't match, that means you have made an error in one of the steps that you followed so far.

V. Docking a single molecule for pose reproduction

Energy minimization

Go to the directory 4.dock and a create a new file (min.in) and enter the command below.

 dock6 -i min.in

Answer the prompted questions using the answers given below or include the following lines in the min.in file at before entering the above command to avoid answering the questions manually.

conformer_search_type                                        rigid
use_internal_energy                                          yes
internal_energy_rep_exp                                      12
internal_energy_cutoff                                       100.0
ligand_atom_file                                             ../1.dockprep/2nnq_lig_withH.mol2
limit_max_ligands                                            no
skip_molecule                                                no
read_mol_solvation                                           no
calculate_rmsd                                               yes
use_rmsd_reference_mol                                       ../1.dockprep/2nnq_lig_withH.mol2
use_database_filter                                          no
orient_ligand                                                no
bump_filter                                                  no
score_molecules                                              yes
contact_score_primary                                        no
contact_score_secondary                                      no
grid_score_primary                                           yes
grid_score_secondary                                         no
grid_score_rep_rad_scale                                     1
grid_score_vdw_scale                                         1
grid_score_es_scale                                          1
grid_score_grid_prefix                                       ../3.boxgrid/grid
multigrid_score_secondary                                    no
dock3.5_score_secondary                                      no
continuous_score_secondary                                   no
footprint_similarity_score_secondary                         no
pharmacophore_score_secondary                                no
descriptor_score_secondary                                   no
gbsa_zou_score_secondary                                     no
gbsa_hawkins_score_secondary                                 no
SASA_score_secondary                                         no
amber_score_secondary                                        no
minimize_ligand                                              yes
simplex_max_iterations                                       1000
simplex_tors_premin_iterations                               0
simplex_max_cycles                                           1
simplex_score_converge                                       0.1
simplex_cycle_converge                                       1.0
simplex_trans_step                                           1.0
simplex_rot_step                                             0.1
simplex_tors_step                                            10.0
simplex_random_seed                                          0
simplex_restraint_min                                        yes
simplex_coefficient_restraint                                10.0
atom_model                                                   all
vdw_defn_file                                                /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn
flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex.defn
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex_drive.tbl
ligand_outfile_prefix                                        2nnq.lig.min
write_orientations                                           no
num_scored_conformers                                        1
rank_ligands                                                 no 

If the process is successful a new file (2nnq.lig.min_scored.mol2) will be generated. You can compare how is it changed from the initial structure by analyzing the RMSD value generated in the file. Visualize the new mol2 file along with receptor and the initial ligand mol2 files using Chimera to see the differences.

2nnq_receptor with the original ligand and the minimized ligand