Difference between revisions of "2008 DOCK tutorial with Gleevec"

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For additional Rizzo Lab tutorial see [[DOCK Tutorials]].
==About DOCK==
==About DOCK==

Revision as of 10:58, 31 January 2011

For additional Rizzo Lab tutorial see DOCK Tutorials.

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.

Installing DOCK

  • Unzip or copy the DOCK distribution to a convenient folder. e.g. /home/sudipto/dock6
  • Go to dock6/install, and run ./configure gnu
  • Type 'make'. A lot of compilation messages should scroll by the screen, with occasional warnings.
  • If everything went well, you should get a message which looks like
Installation of
DOCK v6.2dev
is complete at Mon Feb  4 14:47:28 EST 2008.
  • The DOCK executable should be dock6/bin/dock6
  • Update your .cshrc (or equivalent) file to contain path to DOCK bin directory. This can be done where other program variables have been set.
 ### DOCK ###
 set path = ( $path     $AMS536/dock6/bin)

You need to resource your .cshrc file

source .cshrc

Docking in Gleevec

Gleevec complexed with Tyrosine kinase domain. Image made with Chimera.
Go to the RCSB Protein Databank and search for Gleevec.

You should find the PDB code 1XBB named "Crystal structure of the syk tyrosine kinase domain with Gleevec".

If not, use the link http://www.rcsb.org/pdb/explore.do?structureId=1XBB

Select Download Files in the left window pane. Select "Biological Unit gz". This will give you the biological unit file compressed with gzip. If you can not find it, use the link http://www.rcsb.org/pdb/files/1xbb.pdb1.gz

Download the gzip file and locate it on your desktop. Uncompress the file with the command

gzip -d 1xbb.pdb1.gz

You should now have the file 1xbb.pdb1 Open the file in vi and look at the contents.

It will have a header that looks like this:

HEADER    TRANSFERASE                             30-AUG-04   XXXX              
TITLE    2 GLEEVEC                                                              
COMPND    Tyrosine-protein kinase SYK (E.C.                           
EXPDTA    X-RAY DIFFRACTION                                                     
AUTHOR   4 A.RAMOS, M.RUSSELL, S.K.BURLEY, S.G.BUCHANAN                         
JRNL        AUTH   S.ATWELL, J.M.ADAMS, J.BADGER, M.D.BUCHANAN,                 
JRNL        AUTH 4 B.W.NOLAND, K.POST, K.R.RAJASHANKAR, A.RAMOS,                
JRNL        AUTH 5 M.RUSSELL, S.K.BURLEY, S.G.BUCHANAN                          
JRNL        TITL 2 STRUCTURE OF SPLEEN TYROSINE KINASE                          
JRNL        REF    J.BIOL.CHEM.                  V. 279 55827 2004              
JRNL        REFN   ASTM JBCHA3  US ISSN 1083-351X 

Scroll past the SEQRES entries to see the section describing the ligand

HETSYN     STI STI-571                                                          
FORMUL   2  STI    C29 H31 N7 O                                                 

The first line contains the IUPAC name of the ligand, followed by the molecule 'synonym' used in this file for the ligand. The third line describes the molecular formula of the ligand. Note the number of hydrogen atoms here. It is often useful to verify that the correct number of hydrogen atoms have been added during protonation.

Receptor Preparation with Chimera


Open the file 1xbb.pdb1 with Chimera. Examine the PDB file, observe the binding site and the ligand bound to it.

Now select and delete the ligand Gleevec from the complex.

Go to Actions -> Atoms/Bonds -> Delete to delete the ligand molecule.

Now go to Tools->Structure Editing->Dock Prep. This starts the DockPrep module of Chimera. During DockPrep, it will ask for the histidine protonation. In this select 'Unspecified determined by method'.

After DockPrep has run, save the charged receptor file in the .mol2 format (say 'receptor_charged.mol2). (Go to -> File|Save Mol2)

In order to generate the molecular surface of the receptor, one has to use the 'dms' program. For this the Hydrogens of the receptor needs to be removed.

(Select| Chemistry |Element | H and then Actions| Atoms/Bonds |delete

Now save the mutated receptor in .pdb format (say receptor_noH.pdb). (Go to -> File|Save PDB)

For more details about DockPrep, use http://www.cgl.ucsf.edu/chimera/1.2199/docs/ContributedSoftware/dockprep/dockprep.html

[Sample Prepared Receptor]

Ligand Preparation in Chimera

Gleevec, a popular anti-cancer drug from Novartis. See http://en.wikipedia.org/wiki/Imatinib for more details

Close the previous session in Chimera and reopen the original pdb file '1XBB.pdb1' in Chimera.

Now delete the receptor to get only the ligand. (Select| Structure | Ligand. --> Select | Invert all models --> Actions |Atoms/Bonds |Delete)

To prepare the ligand, select the ligand and add hydrogen and charge using Tools | Structure Editing |Add H and similarly Add Charges. The formal charge of the ligand here is +2. Verify that the hydrogens have been added correctly as in the diagram.

Save the molecule in .mol2 format (say 'ligand_charged.mol2).

Molecular Descriptors for Gleevec:
Rotatable_Bonds:       7
Heavy_Atoms:           37
HBond_donors:          2
HBond_acceptors:       5
Molecular_Weight:      493.61
Log_P:                 3.08

[Sample 1XBB Prepared Ligand]

Generating the Receptor Molecular Surface

This is done by using the 'dms' program and the mutated receptor file with hydrogens stripped off.

/home/sudipto/dms_program receptor_noH.pdb -n -w 1.4 -v -o receptor.ms 

The options associated with the dms program are

-a  	use all atoms, not just amino acids
-d 	change density of points
-g 	send messages to file
-i 	calculate only surface for specified atoms
-n 	calculate normals for surface points
-w 	change probe radius
-v 	verbose
-o 	specify output file name (required)

Generate Spheres

1. Run sphgen on the command line using the sample input file called INSPH. Note, sphgen requires that the input file be calaled INSPH. The output file is always called OUTSPH. Note, if any of the output files (OUTSPH) or any of the temp files (temp1.ms, temp2.sph or temp3.atc,) exist, a core dump will occur. Just delete all these files and start again). SPHGEN takes a few minutes to run.

sphgen -i INSPH -o OUTSPH

Here is a sample output OUTSPH file.

density type = X
reading  rec.ms                                                                             type   R
# of atoms =   2317   # of surf pts =  64657
finding spheres for   rec.ms                                                   
dotlim =     0.000
radmax =    4.000
Minimum radius of acceptable spheres?
output to  rec.sph                                                             
clustering is complete     32  clusters

2. Select spheres in the proximity of the binding site:

sphere_selector takes as input receptor.sph which that was generated by sphgen, a .mol2 file specifying the ligand binding site and the distance (in angstroms) from the ligand.

sphere_selector receptor.sph ligand_charged.mol2 10.0

3. Convert spheres to PDB format using showsphere program so we can visualize the spheres in the context of the receptor and the original ligand

showsphere <showsph.in

The file selected_spheres.pdb (as defined in showsp.in) is generated after running showsphere.

4. View results

Based on the showsph.in file, this will generate a pdb file selected_spheres.pdb.

This section will use command line mode of Chimera. After starting Chimera, use Tools->General Controls->Command Line Adjust the side view to get entire protein in view (Tools->Viewing Controls->Side View

open selected_spheres.pdb
open receptor_charged.mol2
open ligand_charged.mol2
surface #0
~disp #0
surface #1
~disp #1
surface #2
~disp #2
surftrans 50 #0

This will open the spheres, receptor and ligand and turns on the surface display and turns off the atom/bond display. The transparency of the spheres is set to 50%

[Sample Prepared Spheres]

Generate Grid

Generate grid box.

showbox < showbox.in

After running showbox we can view the box that the grid will be placed in.

Start chimera

Open the receptor, spheres from sphgen and the box from showbox.

In Chimera, select

File->Open-> and select your receptor molecule (ie receptor_charged.mol2) File->Open-> and select your box (ie rec_box.pdb) File->Open-> and select your spheres (ie selected_spheres.pdb)

Make the receptor a surface representation. Select->Chain->A Action->Surface->Show Action->Atoms/Bonds->Hide

Select your grid box and enlarge the lines.

Select->Residue->Box Actions->Atoms/Bonds->Wire Width->4

Increase size on the selected spheres. Select->Chain->(No id)->Selected Spheres Action->Atoms/Bonds->Ball+Stick Tools->General Controls->Model Panel

Select the selected_spheres model by clicking once on the name.

Select the 'Attributes' button on the right. Set the ball scale to .5.

The following figure shows the surface representation of the receptor molecule in red. The spheres that occupy the nooks and crannies represent potential sites that heavy atom molecules can potentially occupy.

Rec box sphere.png

One can also run the following commands in command line mode.

open receptor_charged.mol2
open rec_box.pdb
open selected_spheres.pdb
surface #0
~disp #0
linewidth 4 #1
repr bs #2
setattr ballScale .5 #2

Generate grid

grid -i grid.in

This generates grid.nrg (energy scoring file) and grid.bmp (bump grid) files. The grid computation creates an energy grid grid.nrg and a bump grid grid.bmp. This computation should take about 5 mins to finish.

If you need to run grid again, please ensure that grid.bmp and grid.nrg files are removed.

[Sample prepared grids]

Perform Docking

There are two kinds of docking options : Rigid Ligand Docking and Flexible Ligand Docking.

Rigid Ligand Docking In this the ligand is held rigid during orientation.

dock6 -i rigid.in -o rigid.out

This generates the following files:

rigid.out which gives a summary of the run,

rigid_scored.mol2 which gives the MOL2 file containing the geometric coordinates as well as the summary of interaction energies of binding for the docked ligand poses in the order of their grid score. The best pose would have the most favourable grid score. The following shows a sample view of the details of interaction energies generated in the mol2 file.

########## Name:		receptor.pdb
########## RMSD:		1.01108
##########    Grid Score:          -52.555294
##########           vdw:          -37.118080
##########            es:          -15.437213

To visualize the docking result, open rigid_scored.mol2 in Chimera and simultaneously load the receptor.pdb and ligand_charged.mol2 files.

Flexible Ligand Docking

In this type of docking, the ligand is allowed to be flexible by applying an 'anchor and grow' algorithm.

dock6 -i flex.in -o flex.out

This generates the following files:

flex.out giving the summary of the run.

flex_scored.mol2 which gives the geometric coordinates and the summary of the interaction energies of binding for the docked ligand poses in order of their grid score.