2010 DOCK tutorial with Streptavidin

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Revision as of 19:42, 20 February 2010 by Fgorgani (talk | contribs) (Running DOCK)
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About DOCK

DOCK was developed by Irwin D. "Tack" Kuntz, Jr., PhD and colleagues at UCSF. Please see the webpage at UCSF DOCK.

DOCK is a molecular docking program used in drug discovery. This program, given a protein active site and a small molecule, tries to predict the correct binding mode of the small molecule in the active 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.

About Streptavidin & Biotin

Streptavidin is a tetrameric prokaryoke protein that binds the co-enzyme biotin with an extremely high affinity. 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 β-barrel, which has a high affinity as well as a high avidity for biotin. Four identical streptavidin monomers associate to give streptavidin’s 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 neighbouring 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 gluconeogenisis.

Downloading the PDB complex (1DF8)

Download the neuraminidase file from here into your working directory. To get the monomer, download using the blue file arrow icon. Choose biological unit gz under download files folder at the left side. In that way you can downlad a biological gz file. If you download it by using "fetch by ID" in Chimera, you can only download a pdb gz file.

If the pdb gz file is downloaded the monomeric structure of neuraminidase with the bound ligand is seen.

However if the biological gz file is downloaded a tetramer complex of neuraminidase is seen.

When docking, we will only choose a monomer of the whole protein and its ligands instead of the whole tetramer. And it will be painful if you follow the procedure below, use vi to prepare the enzyme and ligand. Because you need to make sure the ligand you extract from the whole pdb files is right the one that match the monomer you've chosen.

Preparing the Enzyme and Ligand in Chimera

To delete the water molecules, etc. go to tools->struc. edit--> dock prep. Check all boxes and click ok. Save as 1DF8.dockprep.mol2 To separate the ligand: select--->residue-->BTN. Then go to actions-->atoms--->delete Save file: 1DF8.receptor.mol2 Select--->chemistry-->element-->H and delete. Save as PDB file 1DF8.receptor.noH.pdb close and open dockprep file again. select protein and delete. Now have ligand only. Save as 1DF8.lig.mol2

Generation of Enzyme Surface, Spheres, and Grid for DOCK

Enzyme Surface

To generate an enzyme surface, first open the receptor pdb file with the hydrogen atoms removed (1DF8.rec.noH.pdb). Next, go to Actions -> Surface -> Show.

Example alt text

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.rec.dms.

The Write DMS algorithm will "roll" a small probe (default radius = 1.4 Angstroms) over the surface of the enzyme and calculate the surface normal at each point. Note that this can also be accomplished with a separate dms program, as described in DOCK tutorials from previous years. DMS (dot molecular surface) files are subsequently used as input for sphgen.


To generate spheres file, we need to use command line program called sphgen. To run the sphgen, we need a input file named INSPH. The content of INSPH is like this


1DF8_receptor.ms is the surface file we got from the previous step. (Change the file name to what you named it before). 1DF8_receptor.sph is the spheres file we want to generate in this step.

Use this command to generate spheres file

sphgen -i INSPH -o OUTSPH

You should get the OUTSPH similar to this

density type = X
reading  1DF8_receptor.dms                                                                  type   R
# of atoms =    881   # of surf pts =  10771
finding spheres for   1DF8_receptor.dms
dotlim =     0.000
radmax =    4.000
Minimum radius of acceptable spheres?
output to  1DF8_receptor.sph
clustering is complete     27  clusters

You can open the spheres file (1DF8_receptor.sph). There are over 700 atoms in this file. However, we're only interested in docking the ligand into the active site. To get better efficiency, we can select those sphere atoms in the active site by using sphere_selector command.

sphere_selector 1DF8_receptor.sph 1DF8_ligand.mol2 10.0

Where 1DF8_ligand is the ligand file. 10.0 is the distance cutoff. You should get a file called selected_spheres.sph. You can open it. The number of atoms is drop down to 47.


Running DOCK

You can run dock with either a rigid or flexible ligand. For either one, you need to creat an input file.

Lets start with a rigid ligand. We need to first make the input file by typing in:

vi rigid.in

The rigid.in file is:

ligand_atom_file                                             1DF8.lig.mol2
limit_max_ligands                                            no
skip_molecule                                                no
read_mol_solvation                                           no
calculate_rmsd                                               yes
use_rmsd_reference_mol                                       no
use_database_filter                                          no
orient_ligand                                                yes
automated_matching                                           yes
receptor_site_file                                           selected_spheres.sph
max_orientations                                             1000
critical_points                                              no
chemical_matching                                            no
use_ligand_spheres                                           no
use_internal_energy                                          yes
internal_energy_rep_exp                                      12
flexible_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                                       grid
dock3.5_score_secondary                                      no
continuous_score_secondary                                   no
gbsa_zou_score_secondary                                     no
gbsa_hawkins_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                                        no
atom_model                                                   all
vdw_defn_file                                                vdw_AMBER_parm99.defn
flex_defn_file                                       /nfs/user03/sudipto/dock6/parameters/flex.defn
flex_drive_file                                 /nfs/user03/sudipto/dock6/parameters/flex_drive.tbl
ligand_outfile_prefix                                        rigid
write_orientations                                           no
num_scored_conformers                                        5000
write_conformations                                          no
cluster_conformations                                        yes
cluster_rmsd_threshold                                       2.0
rank_ligands                                                 no 

Note-The vdw_defn_file was in my directory. The flex_defn_file and flex_drive_file files were in sudiptos directory

Now you can run this file through a .csh. We made ours like this:

vi dock6.rigid.csh
#PBS -l nodes=1:ppn=2
#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/username/1DF8_setup
/nfs/user03/sudipto/dock6/bin/dock6 -i rigid.in -o rigid.out

Docking Results