2019 Covalent docking tutorial 1 with PDB 5VKG

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This tutorial teaches you how to dock a covalently bound drug molecule to a receptor (PDB 5VKG).

Use chimera to visualize your system in this case the pdb 5VKG of NMR structure of human Tsg101 UEV in complex with tenatoprazole. PDB 5VKG consists of 20 different models, but we need only one. Either chose one of the models or ask Lauren to give you the file rcsb104645_model1.pdb

N16 covalently bound to Tsg101 is shown

We are interested in the interaction between S atom in Cys 73 and S atom in the ligand. Those two atoms are covalently bonded according to the experimental paper, but Chimera will not show a covalent linker between these two S. We can create the bond ourself in Chimera, if we need (but we don't actually) and measure the distance between these two atoms. If there is a disulfide bond between them, the distance should be 2.05A. To check the distance we need Shift+Control+left mouse to select two atoms between we want to measure distance.

Tools -> Structure analysis -> Distance. 

You will notice the distance from the two sulfurs is about 2.03 angstroms.

Distance between the ligand S atom and Cys73 S atom is 2.03A.

I. Receptor Preparation

First step is to prepare the receptor. To do this open up the original pdb of the molecule. To prepare the receptor, you need to cut off the covalent linkers to generate the receptor.

Tools -> Structure editing -> Build Structure -> Modify structure - Element S, Bonds 2, Geometry tetrahedral. 

The newly formed receptor is now with a complete cysteine.

Receptor with Neutral cysteine.

Now add hydrogens and charge the protein. To make the cysteine neutral, protons will be added into the system. Protonate with the latest AMBER force field, currently AMBER14SB.

Tools -> Structure editing -> AddH. 
Tools -> Structure editing -> AddCharge.

To prepare it for making a grid, delete the side chain of the covalently bonded cysteine residue. When deleting the side chain of the residue make sure to delete the beta carbon but leave the alpha carbon. That is the receptor that will be used for docking and for the dock grid.

All atoms of the covalently linked cysteine are deleted leaving only the backbone.

II. Ligand Preparation

Open of the original structure. Delete all the protein leaving only the ligand covalently bonded too the cystine, leave the cystine attached to it. The ligand should be binded to the side chain residue all the way up to the beta carbon. Protonate the modified ligand and charge it with AM1BCC. Then delete all of the hydrogens off the beta-carbon. Modify the beta carbon so the name of the residue is D2. Chain or modify the sulfur atom so the name is D1. In VI change the atom type of the beta carbon and the sulfur to Du. The atoms type are being changed to dummy atoms. This is the end of the ligand preparation

Manually edit the mol2 so that the two sulfurs are connected. Short example shown below:

     6 SG         -24.3290   32.6070  -37.1160 S.4       1 CYS    0.0000
    32 S20        -24.6380   34.0060  -37.3390 S.3       1 LIG    0.0000
    1    1    7 1
    2    6   32 1

The @<TRIPOS>ATOM denotes the coordinates of the atoms which should stay unchanged. The @<TRIPOS>BOND denotes the connectivity records in the mol2. You will need to add another connection between the two sulfurs. The first columns is the bond number. The second column is the atom number of the atom seen first (the lowest atom number goes first). The third column is the atom number of the second sulfur (the higher atom number). The fourth and final column is the bond type, 1 being a single bond. Highlighted in bold above shows the modified bond connectivity. The ligand should look connected to the cysteine residue as seen below.

N16 covalently bound to cysteine sidechain

Delete the backbone of the cysteine leaving the beta carbon. Then manually change the atom name of the beta carbon to D2 and change the atom type from C.3 to Du. Change the atom name of the gamma sulfur connected to the beta carbon to D1 and changed the atom type to Du as seen below:

     1 D2        -24.0727   34.9285  -36.1952 Du        1 LIG   -0.0660
     2 D1        -25.1580   36.1840  -35.7130 Du        1 LIG   -0.1111

The ligand should now look like this in chimera:

N16 modified to include the beta carbon and sulfur as dummy atoms.

III. Preparing the sphere files for orienting

The only spheres that will be orientated will be the alpha carbon, beta carbon, and the sulfur on the cysteine sidechain. Isolate the sulfur, alpha and beta carbons and delete everything else. Once that is isolated save it as a pdb or as a mol2. Convert this pdb or mol2 into a sphere file manually. Save the sphere file for orienting when docking.

The three atoms from the cysteine sidechain needed for oritenting: beta carbon, alpha carbon, and sulfur.

Now new spheres will be generated for the box. To accomplish this spheres will be generated ligand file. A sphere file will be made using the coordinates from the ligand file of the heavy atom. Now that the spheres are generated twice once for orientating and the other for the dock/grid. The box will then be generated, once the box is made the grid can be populated in the box. Now that the grid, the ligand, and spheres meant of orientating. You can begin covalent docking.