2022 DOCK tutorial 3 with PDBID 1X70

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System Preparation

Fetching 1X70

Open Chimera and do the following to grab the protein:

 File > Fetch By ID > 1X70
Biological assembly 1 of 1X70.

The first thing to notice is that this is a dimer and the ligand, 715, is not bound at the dimer interface. Thus, one of the monomers is entirely redundant and should be deleted.

 Select > Chain > A
 Actions > Atoms > Delete

Next it is important to remove cofactors, ions, and water molecules not involved in the binding interactions. This can be checked by reading through the paper associated with the PDB.

 Select > Residue > NAG > Actions > Atoms > Delete
 Select > Residue > NDG > Actions > Atoms > Delete
 Select > Residue > HOH > Actions > Atoms > Delete

This will leave us with just the ligand and receptor.

1 monomer of 1X70 with all waters and non standard residues deleted except for the ligand 715, which is colored light blue for clarity.

Receptor Prep

Receptor 1X70 without Hydrogens

Now that we the receptor by itself, we have to clean up the rest of the receptor by adding any missing side chains, dealing with multiple occupancies and mutated residues, and protonating and calculating partial charges.

For 1X70, there are several residues with multiple occupancies.

One way to check for these residues is to grep the number of alpha carbons from the pdb in the command line.

 In the terminal, type: grep -e CA 1X70_noH.pdb
Two potential occupancies shown for CYS649

To clean this up and protonate/charge the receptor:

 Tools > Structure Editing > Dock Prep
Using dock prep to fill in missing atoms/sidechains, add Hydrogens and charges.

Make sure that the protonation makes sense for residues in the active site or coordinated with metals (none here), especially histidines, by checking the paper and chimera for any nitrogens coordinating with metals are not protonated or residues in the active site with differing protonation states. Also make sure that all residues have integral charge (i.e. the charge is an integer). Chimera should give a warning after the dockprep if any residue charges are non-integral.

Ligand Prep

First you have to save the ligand as a separate file. You can do this in Chimera by deleting all of the protein and saving that as a separate file: "715_noH.pdb".

 Select > Residues > 715 > Invert Selection
 Actions > Atoms > delete
 File > save PDB

Now you should just have the ligand.

715 without Hydrogens added

Next Hydrogens and partial atomic charges need to be added and saved as "715_H.mol2". It will ask for the ligands overall charge, which you should verify using chemical knowledge.

 tools > structure editing > addH 
 tools > structure editing > addCharge > Select the correct charge for your ligand, Use AM1-BCC. 
 File > save Mol2
715 with Hydrogens

Surface Generation & Spheres

This section details the generation of sphere files which will be used to describe where you are trying to DOCK to on your protein.

Surface Generation

 Load 1X70 w/o Hydrogens in Chimera
 actions > show > surface

The VDW surface for 1X70 that will be used to generate the sphere files.

Then you will write a DMS (Molecular Surface) File With the surface generated in Chimera: Tools > Structure Editing > Write DMS

Now you should have a DMS file for the next step.

Sphere Generation To generate spheres make the following input file: "INSPH" -

 1X70_dms.dms      #Molecular Surface File 
 R                 #Whether to generate spheres outside of surface (R) or inside (L) 
 X                 #Surface points from the DMS file to use in sphere generation 
 0                 #Minimum radius between spheres
 4.0               #Maximum radius of sphere
 1.4               #Minimum radius of sphere
 1X70_wo_H.sph     #Output sphere file

For more information on sphere generation see: https://dock.compbio.ucsf.edu/DOCK_6/tutorials/sphere_generation/generating_spheres.htm

The .sph file should give you something similar to the following image if you load it up over your protein in Chimera:

The spheres representing the empty space within the protein.

Sphere Selection

Making The Infamous Grid

Making the box

In order to make the grid we first have to determine how big our grid will be. To do this:

 cd 03.grid
   #automatically construct box to enclose spheres [Y/N]
   #extra margin to also be enclosed (angstroms)?
    #(this will be added in all 6 directions)
   #sphere file-
   #cluster number-
   #output filename?

You can also copy these inputs into "showbox.in" (none of the commented lines) and then type "showbox < showbox.in"

Making the grid

Making the following input file "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                             ../01.structures/1X70_rec.mol2
 box_file                                  ./1X70.box.pdb
 vdw_definition_file                       /gpfs/projects/AMS536/zzz.programs/dock6.9_release/parameters/vdw_AMBER_parm99.defn
 score_grid_prefix                         grid

Now generate the grid. It should take several minutes.

 grid -i grid.in -o gridinfo.out

Once it is done, vi into gridinfo.out and make sure the charges are all integer and that the calculation finished. If they are not, that likely means there is something wrong with your 1X70_H.mol2.

You should get two files:

 grid.nrg (energy grid)
 grid.bmp (bump grid)

Try loading them into chimera over your protein.

The energy grid for 1X70.
The bump grid for 1X70.

Energy Minimization for the ligand

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Docking & Virtual Screening

Rigid Docking

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Fixed Anchor Docking

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Flexible Docking

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