After selecting Tools->Ligand docking, the following procedures will take place.

Selecting a Small Molecule for Docking

choose a small molecule for docking by clicking on it.

You can make it by using both the 3D structure (Figure 25) (b) and the 1D structure (a).

Figure 25 - Small Molecule Selection

After selecting a ligand, the viewer zooms in on the ligand and shows the molecule in sticks.

Press the Next button in the Wizard (c) to proceed with the following steps.

Setting the Protonation State and Adding Explicit Hydrogen atoms to the Ligand

BACKGROUND: Most sources of 3D structures provide molecules only with heavy atoms and without hydrogen atoms. There is also no information on bond types (single, double etc.) and protonation state (adding or extracting a hydrogen atom depending on the pH and the chemical group). But it is essential for Quantum's calculations to have the right number of hydrogen atoms in molecules. This procedure helps to do this.

The small molecule, which was selected in the previous step, is added as a separate object under the name "ligand" (Figure 26) (b).

The notice on the display (a) informs you that you should fix hydrogen atoms and bond types.

Figure 26 - The Ligand

You have three choices here:

·   Go to the next step if you think the ligand has the right number of hydrogen atoms.

·   Use the Build Model option (c) from Wizard to set the protonation state and add hydrogens by using Quantum's algorithms.

·   Use Builder (Figure 27) to manually set bond types and add/remove hydrogen atoms.

Figure 27 - Builder

Use options (a) to deliver hydrogens and (b) to change the number of bonds. To do this , you should use the atom selection (left click) and the bond's selections (right click). You can find more information on working with Builder in the section  Moving and Building Molecules.

Note: We recommend that you use the Build Model option. Build Model analyses the geometry of the molecule - bond lengths and bond and dihedral angles - and adds missing hydrogen atoms. The Build Model procedure also sets the right protonation states (Figure 28).

Figure 28 - Fixing the Ligand

Note: However, you should take into account that in some cases that molecules from the Protein Data Bank do not have the right geometry, and you have to fix them without the assistance of the Build Model procedure or, you have to at least manually correct them after Build Model.

Press the Next button (a).

Building a Grid box in the Active Site

BACKGROUND: Whenever the ligand position inside the active site is not known, the program uses optimization to minimize the binding free energy using Quantum's advanced energy evaluation algorithm. The recommended way to speedup the calculation is to employ precalculated potentials stored in the grids. A grid is a 3D box (gridbox) characterized by three orthogonal directions and its size.

In order to construct a grid box, the following requirements should be met:

·   All active site atoms should be covered by a grid box, or at least all important chemical groups of the active site should lie inside of the grid box.

·   Do not make a grid box too large. A larger grid makes the grid calculation last longer, and it takes more memory to store the grid values in the computer's RAM. The time of the grids' calculations is proportional to the grid box volume.

Note: If you make each side of the grid box two times longer, then the calculation time increases 8 (2x2x2) times. The same is true for the volume of the memory required for using grids.

·   We recommend that the grid box volume does not exceed 20x20x20 A3. For instance, it can be 40x20x10 A3 or 5x40x40 A3 and so on.

First, select the center of the grid box by selecting any atom that lies approximately in the middle of the active site.

After clicking, a cube with the dimensions 20x20x20 A3 will appear.

Now you have to adjust the box position and size.

·   The grid box has axes OX, OY and OZ, which are displayed on the screen.

·   If you decide to change the size of one of the axes, you should use Grid box Wizard (Figure 29).

Figure 29 - Grid box Wizard

·   Just click on the size button and choose the size. The grid box will automatically be changed.

Note: The default size is 20A. The following options are available: 5A, 10A, 15A, 20A ( default) and 25A.

Selecting Essential Metal Ions and Hetatoms in the Active Site

BACKGROUND: The small molecule can interact with metal ions and hetatoms in the active site. Correct energy calculation should involve all of the important structures within the active site. After the grid box is defined, all structures within it will be renamed (Figure 30) and shown.

Figure 30 - Active Site Structures
 

We have three new objects here: protein, metal and hetatom.

·                   We recommend that you do not include water molecules in modeling since Quantum has its own model of water.

You can remove any structure from the list.

Increase the grid box size to add a structure if necessary.

Adding hydrogen atoms to the protein and to the hetatoms

You have three choices here:

·   Go to the next step - "Dock the Molecule"- if you think the protein and all other structures in the active site have the right number of hydrogen atoms.

·   Use the Build Model option from the Wizard to set the protonation state and add hydrogens by using Quantum's algorithms. We recommend this option.

·   Use Builder (Figure 27) to manually set bond types and add/remove hydrogen atoms.

Figure 31 - Docking Wizard

Choose the option regarding Protein Flexibility (Figure 31), which is off by default.

If this option is OFF, then the protein is treated as a rigid structure. If it is ON, then full Protein flexibility will be taken into account.

Note: Protein flexibility capability depends on the license you purchased. This manual describes all possible functions, and some of them may not be accessible in your installation.

Press the Dock the Molecule button to start modeling.

Calculations and Results

All stages of the process are displayed on the Progress Bar and in the Information Panel.

When the calculation is finished, you can see the results in a window that will appear instead of the information panel (Figure 32).

The window includes:

·   Name - name of ligand

·   IC50 uMol/L - IC50 value

·   E bind, kJ/mol - free binding energy

·   E es, kJ/mol - electrostatic and solvation energy

·   E vdw, kJ/mol - short range electrostatic and exchange and Van der Waals energies

·   TdS, kJ/mol - entropy contribution

·   E tor, kJ/mol - ligand internal energy change

·   Charge, Mass, Flex.bonds - total charge, mass and number of flexible bonds of the ligand

·   RMSD, A - root mean square distance between the initial and final positions

Note: Free binding energy is equal to the sum of all listed contributions (E es, E vdw, TdS and E tor). In our calculations, IC50 is 5.82 x (E bind).

Figure 32 - Results

You can compare the initial and final positions of the ligand by using Viewer. The procedure will create the object ligand_pos with final coordinates.