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Tutorial Molecular Docking: Benzamidine inhibitor docked into Beta Trypsin

In this tutorial you will learn how to set up docking calculations. You will dock the inhibitor benzamidine into to the serine protease beta trypsin. Docking calculations attempt to place Ligands into Binding Sites. Before you can dock a molecule, you first need to define the atoms that make up the Ligand (drug, inhibitor, etc.) and the Binding Site on the protein where the drug binds. The structure we will use is from the Brookhaven Protein Databank and already contains a co-crystallized benzamidine. We will make a copy of the inhibitor and dock it to the protein and compare the docked structure with the x-ray structure.

1. Open the structure file.

Open the protein databank file, 3ptb.ent located in the Tutorials/Docking/Beta Trypsin folder of your ArgusLab installation. This is a PDB file so make sure you select the correct file type in the file open dialog.

Make sure the Molecule Tree View tool is visible. Either select Tools/Molecule Tree View menu option or click the toolbar to toggle the Tree View.

button on the

Expand the tree view of 3ptb and open up the Residues/Misc folder to show the Benzamidine residue. You should see something like this:

2. Make the Ligand and Binding Site groups.

Left-click on "1 BEN" in the Tree View to select the benzamidine residue. It should appear yellow.

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Select the Edit/Hide Unselected menu option or use the shortcut (Ctrl+U) to hide all atoms that are not selected. The only atoms showing on the screen should be the benzamidine molecule. Center benzamidine in the window by selecting the View/Center Molecule In Window menu option or pressing "Alt+C" shortcut, or pressing the button on the toolbar. button on the toolbar. You

While benzamidine is selected, add hydrogens to it by holding down the "Shift" key and pressing the should see something like this:

Right-click on "1 BEN" in the Tree View and select the "Make a Ligand Group from this Residue" option. ArgusLab will construct a group underneath the Groups folder with the same name "1 BEN" that is of type = Ligand. Left-click on either the "1 BEN" residue in the Residues/Misc folder or the "1 BEN" group in the Groups folder. Either action will select the atoms of the Ligand on the screen. Copy (Ctrl+C) & Paste (Ctrl+V) the selected residue. Look in the Residues/Misc folder in the Tree View tool and you will see a new highlighted residue with a name like "810 BEN". Make a Ligand group of this new residue in the same manner you did above. Right-click on "810 Ben" and select "Make a Ligand Group from this Residue". You should now have two Ligands in the Groups folder named "1 BEN" and "2 BEN". Rename the two Ligand groups: Right-click on "1 BEN" in the Groups folder and select "Modify Group..." option. In the Modify Group Dialog box, type in a new name. Let's call it "ligand-xray". Make sure you don't change the Groups type. It should be a Ligand. Do the same for "2 BEN", but name it "ligand".

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To more easily distinguish the two ligand in the graphics window, you can do a number of different things. One would be to choose the View/Color By Molecule option. Another would be to change the rendering style of one of the ligands. I generally prefer the latter. Right-click on the Group named "ligand" and select "Set Render Mode" and choose the "Cylinder med" option. Note, the pasted Group (named "ligand") does not need to be in any special place on the screen. It could be anywhere you wish. It will be moved during the docking. Make the binding site for the ligand-xray group. The easy way is to right-click on the ligand-xray group in the Groups folder and select the "Make a BindingSite Group for this Group" menu option. This will generate a BindingSite that consists of all residues that have at least one atom within 3.5 Angstroms from any atom in the ligand-xray group. This generally gives a good representation of the important residues in the binding pocket for a protein target. Center the molecule again. You should see something like this:

3. Dock the ligand into the binding site

CAVEAT!! When a Ligand group is docking, it will ignore all other atoms that belong to any other Ligand groups that are present. Thus, you can dock multiple ligands into the same structure and see the final results overlaid on each other. However, if you forget to create a Ligand group of a molecule that already occupies the binding site, it will adversely affect the docking results.

Bring up the Dock Settings dialog box by selecting the Calculation/Dock a Ligand... menu option or clicking on the toolbar. Here is what the Dock Settings dialog box looks like:

button on the

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Select the ligand to dock in the "Ligand" drop-box. Make sure to select the group named "ligand" group and NOT the "ligand-xray" group. Click on the "Calculate Size" button and a docking box tailored to the binding site will be make and shown on the screen. Make sure "ArgusDock" is the docking engine, the Calculation type = Dock, and the Ligand is Flexible. Click the "Start" button and the docking calculation will begin. Notice the messages that appear in the status line at the bottom of the molecule window. You should see ArgusLab first generate the scoring grids used during the docking (this should take about 5-10 seconds), then the various search phases will occur, and finally the candidate poses should be processed and the calculation is done. Only the coordinates of the group named "ligand" should have been modified. The position of the two Ligand groups should be reasonably close together.

4. Analyzing the results

Let's see how close the docked structure is to the x-ray structure. In the Tree View tool, select both the ligand and ligand-xray groups by holding down the "Ctrl" key and left-clicking on both groups. Both groups should be highlighted on the screen. Now, right-click on the "Groups" folder tab in the Tree View and select "Calc RMSD position between two similar Groups". An information dialog will appear on the screen giving the root mean square deviation (RMSD) difference in the coordinates of the two ligands. The value should be about 0.5 Angstroms. There are two sources of information about the docking calculation: The log file and the Tree View tool. Expand the ArgusDock calculation in the Calculations folder in the Tree View. You will see a listing of the docking settings and several "poses" starting with the most stable as pose 1. (A pose is merely the name given to the coordinates of a docked structure). Any property in an ArgusLab calculation that has a Bunsen burner icon is a renderable property. Right click on pose #2 and select the "Display" option. The coordinates of ligand will change to this pose and its score will be displayed on the screen. You can test the RMSD difference between the x-ray and docked ligand at this point in the same fashion as you did above. The screen should look like this:

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Right-click on Pose 2 and select "Hide" or "Hide all Props" to turn off display of this particular pose. The display will return to the default display of the lowest energy (most stable) pose. To see the log file, select the Edit/Latest output file... menu option or press the button on the toolbar. The text log file will come up in Notepad. It gives more detail about the atom typing, grid details, and status messages during the docking that may be of some use. This file is overwritten with every new calculation. Finally, if you want to save all the work you have done, you will need to save this molecule to an ArgusLab .agl file. Select the File/Save As... menu option and choose the ArgusLab file type and save it. The docking results will be saved and can be studied at a later date if you like.

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