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2023.04.11.6OIM
This is a work in progress...
>> mkdir 001_structure 002_surface_spheres 003_gridbox 004_dock 005_enrichment
Download the 6OIM PDB file from the RCSB Protein Data Bank by going to https://rcsb.org/structure/6OIM and selecting "PDB Format" in the Download Files drop-down menu in the top right. Save the file in the 001_structure directory. Alternatively, while in the 001_structure directory, run the following command in the terminal:
>> wget https://files.rcsb.org/download/7RPZ.pdb --no-check-certificate
Open 6OIM.pdb in Chimera.
Isolate the receptor and cofactors
Select the ligand and water molecule by typing select #0: MOV, HOH in the command line. The selected atoms should be highlighted in green. Delete the selected atoms by selecting Actions -> Atoms/Bonds -> delete in the menu bar, or by typing delete selected in the command line.
Add hydrogens and charges
Chimera misplaces a hydrogen on GDP when adding hydrogens, the misplaced hydrogen must be corrected before adding charges. Add hydrogens by selecting Tools -> Structure Editing -> AddH in the menu bar, select "Unspecified (determined by method)" and click OK. Select the misplaced hydrogen (H3) and delete it. Select the nitrogen that should be protonated (N1) and select Tools -> Structure Editing -> Build Structure in the menu bar. In the "Build Structure" dialog box that opens, change the mode from "Start Structure" to "Modify Structure" at the top. After "Modify Structure" is selected, under "Change selected atoms to...", change the element to nitrogen and change the number of bonds to 3. Under "Residue Name", select "Change the modified residue's name to" and replace UNK with GDP. Apply the changes and close the dialog box. The newly added hydrogen is named incorrectly, select H1 and reopen Build Structure. Switch to "Modify Structure", change the element to hydrogen, set the number of bonds to 1, set the atom name to HN1, and apply. After renaming, the bond length between the hydrogen and nitrogen should change.
[ image ] [ image ]
Now, add charge by selecting Tools -> Structure Editing -> Add Charge in the menu bar and clicking OK in the dialog boxes that open. The magnesium ion should have a net charge of +2.
[ image ]
Save the structure as kras_cof.mol2 and close the session.
Reopen 6OIM.pdb.
Isolate the ligand
Select the ligand by typing select #0: MOV in the command line. Invert the selection by typing select invert in the command line. Delete the selected receptor, cofactors, and water molecules, leaving only the ligand.
Run Dock Prep
Run Dock Prep by first selecting Tools -> Surface/Binding Analysis -> Dock Prep in the menu bar and clicking OK in the "Dock Prep" dialog box that opens. Select "Unspecified (determined by method)" and click OK in the "Add Hydrogens for Dock Prep" dialog box that opens to add hydrogens. Leave the default selections and click OK in the "Assign Charges for Dock Prep" and the "Specify Net Charges" dialog boxes to assign partial charges. The ligand should have a net charge of [?].
[ image ]
Save as amg510.mol2 and close the session.
Generating DMS
Open kras_cof.mol2.
Run surface calculation by selecting Actions -> Surface -> Show in the menu bar or by typing surf #0 in the command line.
[ image ]
Save the surface as a DMS file by selecting Tools -> Structure Editing -> Write DMS in the menu bar. Save as kras_cof.dms.
Creating surface spheres
Change directories to the 002_surface_spheres directory:
>> cd ../002_surface_spheres
Create a file called INSPH by running the command:
>> vim INSPH
Format the file as follows:
../001_structure/kras_cof.dms
R
X
0.0
4.0
1.4
6OIM.sph
This file will be the input file for the sphere generation program, sphgen. The first line of the file specifies the input DMS file. In the second line, R indicates that spheres should be generated outside the surface. In the third line, X indicates that all surface points should be used. The fourth line specifies the acceptable steric clash in angstroms, which in this case is set to 0.0 or no steric clash allowed. The fifth and sixth lines specify the maximum and minimum sphere radii in angstroms, respectively. The last line specifies the output sphere file name.
Run sphgen to generate spheres based on the parameters given in the INSPH file:
>> ${dock6_path}/bin/sphgen -i INSPH -o OUTSPH
[ dock6_path is a variable (placeholder) for a path like: /home/baliuste/zzz.github/dock6_2019_06_14 ]
The surface spheres generated by sphgen can be visualized in Chimera. In a new Chimera session open 7RPZ.pdb and show the surface. In the same session, open the output sphere file (6OIM.sph) created by sphgen to visualize the surface spheres.
[ image ] [ image ]
Selecting relevant spheres
To select spheres based on the distance from the ligand, run the command:
${dock6_path}/bin/sphere_selector 6OIM.sph ../001_structure/mrtx1133.mol2 3.0
The first two arguments passed to sphere_selector specify the surface sphere and ligand files, respectively. The last argument, 3.0, specifies that only spheres within 3.0 angstroms of the ligand should be selected.
The selected_spheres.sph file generated by sphere_selector can be visualized in Chimera.
[ image ] [ image ]
Change directories to the 003_gridbox directory.
Box generation
Create an input file for box generation called showbox.in:
>> vim showbox.in
Format the file as follows:
Y
8.0
../002_surface_spheres/selected_spheres.sph
1
6OIM.box.pdb
In the first line of the file, Y (Yes) indicates that showbox should create a box. The second line specifies the distance, in angstroms, between the spheres and the box borders. The third line specifies the input sphere file. The fourth line indicates the cluster number. The last line specifies the output box file name.
Generate a box based on the parameters given in showbox.in by running showbox as follows:
>> ${dock6_path}/bin/showbox < showbox.in
The 6OIM.box.pdb file generated by showbox can be visualized in Chimera.
[ image ]
Grid generation
Create the input file for grid generation called grid.in:
>> vim grid.in
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 9
distance_dielectric yes
dielectric_factor 4
bump_filter yes
bump_overlap 0.75
receptor_file ../001_structure/kras_cof.mol2
box_file 6OIM.box.pdb
vdw_definition_file ${dock6_path}/parameters/vdw_AMBER_parm99.defn
score_grid_prefix grid
Generate grids by running:
>> ${dock6_path}/bin/grid -i grid.in -o grid.out