Small Molecule Docking using Autodock4 In this exercise you will dock a small molecule (ligand) to protein (native) and analyze different docking poses. In the first part we cheat a bit and take a protein with a ligand bound, remove the ligand and try docking it back to the protein. In the second part, we use the same ligand but now we will perform the docking to model of a protein. Open a terminal (To left corner: Applications-­‐>System Tools -­‐>Terminal) In the terminal do the following command: source /edu/bjowa51/Public/docking
It will open AutoDockTools, which is a Graphical User Interface that can be used to generate input files to Autodock. Preparations: •
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Navigate to http://www.ifm.liu.se/edu/coursescms/nked15/labs/ the download the 3D coordinates for the structure we will work with today native.pdb, native_ligand.pdb,
ligand.pdb, and cataglyphis_fabp.pdb and store them somewhere so you can find them when you need them. Before we can launch the docking program there are three steps that needs to setup: 1. Setup the ligand, defines the degrees of freedom for the ligand, and the translation-­‐
rotation center. 2. Setup the receptor grid, used to pre-­‐calculate interaction energies with all atom types of the ligand. 3. Setup the docking run, define how long to run etc. Assignment 1: Dock the ligand back to the protein it came from 1.1 Setup ligand: •
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In the AutoDockTools menu, open ligand.pdb: {ligand} -­‐-­‐> {input} -­‐-­‐> {open}, choose to show *.pdb (PDB file), default is *.pdbqt which a Autodock internal format, and open the coordinates for the ligand which you downloaded before (ligand.pdb). Define the atom that will be use as center for translating and rotating the ligand: {ligand} -­‐-­‐> {torsion tree} -­‐-­‐> {detect root} Finally, save your ligand: {ligand} -­‐-­‐> {output} -­‐-­‐> {save} 1.2. Setup receptor grid: •
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Load the receptor molecule: {Grid} -­‐-­‐> {macromolecule} -­‐-­‐> {open}, choose to load a PDB file (*.pdb) and open native.pdb. You will be asked to save the resulting file. Save it as “native.pdbqt”. Choose which ligand to dock into the receptor: {Grid} -­‐-­‐> {set map types} -­‐-­‐> {choose ligand} and choose “ligand” in the list. Define a grid-­‐box (the area which will be searched for a ligand position): {Grid} -­‐-­‐> {Grid Box} o In the menu of the dialog box, click on {Center} then on select {center on macromolecule} o Adjust the box if needed (not needed) and close the window through: {File} -­‐-­‐> {close saving current} Store the grid: {Grid} -­‐-­‐> {output} -­‐-­‐> {save gpf} (choose a name, for example “native.gpf”) Finally, compute grid maps by running: autogrid4 -p native.gpf in the terminal. 1.3. Setup the docking run: •
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Specify recptor: {Docking} -­‐-­‐> {macromolecules} -­‐-­‐> {set Rigid Filename} and choose “native.pdbqt” Choose ligand: {Docking} -­‐-­‐> {Ligand} -­‐-­‐> {choose} -­‐-­‐> (click on your ligand) -­‐> {select ligand}-­‐-­‐> {accept} Setup search parameters: {Docking} -­‐-­‐> {Search parameters} -­‐-­‐> {genetic algorithms}. Change to “short” and accept. Save your settings: {Docking} -­‐-­‐> {Output} -­‐-­‐> { Genetic Algorithms } and enter a name, for example “native_docking.dpf” 1.4. Docking •
Finally, in terminal run autodock:
autodock4 –p native_docking.dpf –l native_docking.out
This will take approximately 4 minutes 1.5. Analysis: •
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Open pymol in the terminal: pymol native.pdb ligand.pdb native_docking.out & This will load the native.pdb ligand.pdb and the docking poses into pymol. If you display native_docking in sticks, you can toggle through the lowest scoring energy poses with “play” button in pymol. Examine the different docking poses in pymol compare with the energy and cluster size found in when opening native_docking.out in a text editor. o Is there any major difference between the conformations (position in pymol as well as energy and cluster size from the grep command)? o Based on the cluster size and energy, does the best position seem like a good one? Assignment 2: Dock the ligand back to a protein model Now you will be slightly more realistic and dock the same ligand to a model of a protein. 2.1. Setup ligand: •
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If you setup the ligand in assignment 1 you don’t need to redo it here since we are using exactly the same ligand. You can go to 2.2 Setup receptor grid. In the AutoDockTools menu, open ligand.pdb: {ligand} -­‐-­‐> {input} -­‐-­‐> {open}, choose to show *.pdb (PDB file), default is *.pdbqt which a Autodock internal format, and open the coordinates for the ligand which you downloaded before (ligand.pdb). Define the atom that will be use as center for translating and rotating the ligand: {ligand} -­‐-­‐> {torsion tree} -­‐-­‐> {detect root} Finally, save your ligand: {ligand} -­‐-­‐> {output} -­‐-­‐> {save} 2.2 Setup receptor grid: •
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Load the receptor molecule: {Grid} -­‐-­‐> {macromolecule} -­‐-­‐> {open}, choose to load a PDB file (*.pdb) and open cataglyphis_fabp.pdb. You will be asked to save the resulting file. Save it as “cataglyphis_fabp.pdbqt”. Choose which ligand to dock into the receptor: {Grid} -­‐-­‐> {set map types} -­‐-­‐> {choose ligand} and choose “ligand” in the list. Define a grid-­‐box (the area which will be searched for a ligand position): {Grid} -­‐-­‐> {Grid Box} o In the menu of the dialog box, click on {Center} then on select {center on macromolecule} o Adjust the box if needed (not needed) and close the window through: {File} -­‐-­‐> {close saving current} Store the grid: {Grid} -­‐-­‐> {output} -­‐-­‐> {save gpf} (choose a name, for example “cataglyphis_fabp.gpf”) Finally, compute grid maps by running: autogrid4 -p cataglyphis_fabp.gpf in the terminal. •
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2.3. Setup the docking run: •
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Specify recptor: {Docking} -­‐-­‐> {macromolecules} -­‐-­‐> {set Rigid Filename} and choose “cataglyphis_fabp.pdbqt” Choose ligand: {Docking} -­‐-­‐> {Ligand} -­‐-­‐> {choose} -­‐-­‐> (click on your ligand) -­‐> {select ligand}-­‐-­‐> {accept} Setup search parameters: {Docking} -­‐-­‐> {Search parameters} -­‐-­‐> {genetic algorithms}. Change to “short” and accept. Save your settings: {Docking} -­‐-­‐> {Output} -­‐-­‐> { Genetic Algorithms } and enter a name, for example “cataglyphis_fabp_docking.dpf” 2.4. Docking •
Finally, in terminal run autodock:
autodock4 –p cataglyphis_fabp_docking.dpf –l cataglyphis_fabp_docking.out
This will take approximately 4 minutes 2.5. Analysis: •
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Open pymol in the terminal: pymol native_ligand.pdb cataglyphis_fabp.pdb cataglyphis_fabp_docking.out & This will load the native_ligand.pdb and the docking poses into pymol. cataglyphis_fabp.pdb is actually a homology model made using native.pdb as a template, so you might expect that the ligand will have a similar binding mode. If you use the “align” function in pymol you can superimpose the native_ligand.pdb on top of cataglyphis_fabp.pdb and get a rough idea on the “correct” binding pose. If you display cataglyphis_fabp_docking in sticks, you can toggle through the lowest scoring energy poses with “play” button in pymol. Examine the different docking poses in pymol compare with the energy and cluster size found in when opening cataglyphis_fabp_docking.out in a text editor. o Is there any major difference between the conformations o Based on the cluster size and energy, does the best position seem like a good one?