Section - 1: Molecule Sketching

Go Back

[A.] To understand and draw molecule sketching , SMILE file format and calculating properties

Introduction

Biologically active compound act as a drug.these software is helpful for sketching molecule. Smiles file format is The Simplified Molecular Input Line Entry Specification (SMILES) a line notation for molecules. SMILES strings include connectivity but do not include 2D or 3D coordinates.

Hydrogen atoms are not represented. Other atoms are represented by their element symbols B, C, N, O, F, P, S, Cl, Br, and I. The symbol "=" represents double bonds and "#" represents triple bonds. Branching is indicated by (). Rings are indicated by pairs of digits.

Name      Formula      SMILES String
Methane      CH4        C
Ethanol      C2H6O        CCO
Benzene      C6H6        C1=CC=CC=C1 or c1ccccc1
Ethylene      C2H4        C=C

STEPS:

• Open the site
  http://www.molinspiration.com


• click on free on-line cheminformatics services


• Use the icon | for single carbon || for double bond


• Red cross is eraser


• Sketch molecule aspirin.


• For it Open site
  http://www.ncbi.nlm.nih.gov/sites/entrez?db=pccompound


• search for aspirin


• For smiles format take it from pubchem canonical smiles


• paste smiles format CC(=O)OC1=CC=CC=C1C(=O)O in paste smiles here.


• click on calculate properties and predict bioactivity


• You will get output predict bioactivity



GPCR ligand	-0.66
Ion channel modulator	-0.91
Kinase inhibitor	-0.49
Nuclear receptor ligand	-1.23



• You will get output for calculate properties.




miLogP	1.434
TPSA	63.604
natoms	13
MW	180.159
nON	4
nOHNH	1
nviolations	0
nrotb	3
volume	155.574

Section - 2

Go Back

[A.] Protein-Ligand Docking in AutoDock

Starting Autodock

open directory where MGL tool is installed

(home/MGLtool/bin) and run 'adt' in terminal

It opens AutoDock Tool GUI.

Protein - Ligand Docking in AutoDock

For this you should have a protein molecule 3d-structure file and a ligand 3d-structure file.

Part - A

Open Protein Molecule in viewer

( File -> Read Molecule)

browse and open protein structure file

Visualize Protein Structure

Use various option from top menu and visualize protein structure. Preparing PDB file

PDB files are not perfect every time when it is opened very first time in ADT. There are some potential problems such as missing atoms, chain breaks, added water or other hetero atoms etc. You have to correct these problems from PDB file by adding or removing required elements.

• Removing 'Water' molecule or hetero atoms from PDB-
  
  For this, go to
  Select -> Select from string
  This opens a box contains four option to select molecule, chain, residue or atom.
  Enter 'HOH*' in residue box and click on 'Add'. It select all water molecules of protein structure.
  You can enter other residue name(s) for their selection. For multiple selection enter name and number of all residues seperated with column containing no spaces. (example: ARG1,TYR39,PRO12)
  
  You can choose other option too, to select content at their level.
  After selection of water molecules in structure, go to
  
  Edit -> Delete -> Delete Selected Atoms
  
  It will ask for confirmation. Continue to it.



• Adding 'Hydrogen' to set noBond Order to molecule
  
  Edit -> Hydrogen -> Add
  
  It will ask for add hydrogen, choose All Hydrogens with noBondOrder and click OK.



• Save as PDB file
  To keep original file safe, save this modified protein structure to a new pdb file.
  
  File -> Save -> WritePDB
  
  Select write options and click to OK.

Part B

Preparing Ligand file

Now this is time to open ligand molecule into ADT and prepare it for docking. For this go to
Ligand -> Input -> Open

Browse and open ligand molecule.
This ligand should have torsional bonds and partial atomic charges with it so can assosiate with a better position in protein. So there is a rigid part in ligand known as 'root' which keeps hold torsional bonds and flexible parts known as branches emanate from root, which shows torsions. A ligand must have both polar and non-polar hydrogens. When you open a ligand file in ADT, it automatically computes Gasteiger charges(partial charges) and rotatable bonds, and if no charges present, ADT tries to add charges in ligand and also adds a TORSDOF (Torsion Degree of Freedom) value.
This ligand file is written in PDBQT format where Q and T stand for partial charges and torsion angles. Means this file has additional information of partial charges and torsional angles.
Detect Root and Torsions for ligand
For this, go to

Ligand -> Torsion Tree -> Detect Root

ADT detects for the finest root in ligand and show a ball shaped root marker.
To show/hide this root marker
Ligand -> Torsion Tree -> Show/Hide Root Marker

Choose torsions in ligand
To view and choose torsional bonds in ligand, go to

Ligand -> Torsoin Tree -> Choose Torsions

A 'Torsion Count box' will appear showing information about number of rotatable bonds. There are three types of bonds represented by three different colors-

• Green - Rotatable bonds*


• Magenta - Non-Rotatable bonds*


• Red - Unrotatable bonds

(*These rotatable and non-rotatable bonds can be converted into each other by clicking on them while honding 'Shift' key.)
So choose desired rotatable/torsional bonds in ligand and click on 'done' on Torsion Count box.
To select only a number of torsional bonds, go to

Ligand -> Torsion Tree -> Set Number of Torsions

Checking partial atomic charges on ligand
For this, go to

Edit -> Charges -> Compute Gasteiger

Save ligand to file
Ligand -> Output -> Save as PDBQT
Write ligand name with extension 'pdbqt' added (example: ligand.pdbqt) and save it to default directoty.

Part C

Generating Grid Parameter File (GPF)

A grid parameter file is that which comptes records about the receptor. These records are in form of various maps such as locations, electrostatic, potential energy etc. This GPF file tells autodock these information of receptor that where will the docking be performed. Steps to generating GPF file are as follow-

Grid -> Macromolecule -> Open

Browse and open that 'protein.pdbqt' file. This will open rigid part of receptor protein molecule.
Now ligand and flexible residue file will be selected. For this, go to

Grid -> Set Map types -> Choose Ligand

This will open select molecule box, select ligand molecule from there.

Grid -> Set Map types -> Open protein.pdbqt

Browse and open 'protein.pdbqt' file. This will open flexible residue part of receptor protein molecule.

Setting up grid box

A grid box is to be set where the docking will be occured. This grid box is a minimum volume containing only pocket portion of receptor i.e. flexible residue part. For this, go to

Grid -> Grid Box

This will open a Grid Option Box and a box will also appear in visualizer. This Grid Option Box contains two sections, the above one is to set volume of this grid box in X,Y and Z axis. The below section is to move this grid box in X,Y and Z axis. Set up this grid box according to your docking area and after that -

Grid Option Box -> File -> Close saving current

Now this grid is saved as a file. For this, go to

Grid -> Output -> Save GPF

Save gpf file in default directory with a name following '.gpf' extension (example: protein.gpf)

Now this is time to run this GPF file so that grid log file and required maps files can be generated. For this, go to

Run -> Run AutoGrid

This will open Run AutoGrid box. Here we set up parameterd to run autogrid.

• Program Pathname - Browse and select path of autogrid bin file.


• Parameter file name - Browse default directory and open GPF file.


• Log filename - When you browse for GPF file, after that it automatically sets path and log filename follwing extension '.glg'.


• Set Nice Level to 20 and Launch AutoGrid.

After a few seconds, this process will complete telling whether succesfull or not.

Part D

Generating Docking parameter File (DPF)

A DPF file is that which contains records of all selected parameters and desired map file's informations that will help autodock to compute position of ligand into receptor protein molecule. Means it is used to perform final protein-ligand docking.

Following steps are taken for docking-

• Set rigid part of receptor molecule
  
  Docking -> Macromolecule -> Set Rigid Filename


• Set flexible residues of receptor
  
  Docking -> Macromolecule -> Set Flexible Residue Filename
  


• Set ligand
  
  Ligand -> Choose (Select ligand and click on select ligand)
  


• Set energy evolution parameters
  
  Search Parameter -> Genetic Algorithm
  Set Maximum number of evals to short, so the process can be completed in less time.
  


• Docking Parameters -> Accept



• Save Docking parameter file (DPF)
  
  Output -> Lamarckian GA
  Browse the default directory and save 'ligand.dpf' file.
  

After set up all parameters for receptor and ligand, finally docking will be performed. For this, go to

Run -> Run AutoDock

This will open Run AutoDock box. Here we set up parameterd to run autodock.

• Program Pathname - Browse and select path of autodock bin file.


• Parameter file name - Browse default directory and open DPF file.


• Log filename - When you browse for DPF file, after that it automatically sets path and log filename follwing extension '.dlg'.


• Set Nice Level to 20 and Launch AutoDock.

After a few seconds, this process will complete telling whether succesfull or not. This will also create a Docking Log File which contains final results of protein-ligand docking.

Part E

Visualization of Protein-Ligand Docking

Now we can visualize performed docking in ADT visualizer. For this, go to

• Analyze -> Docking -> Open
  
  Browse to default directory and open docking log file (ligand.dlg)


• Macromolecule -> Choose (or can Open)
  
  Select 'protein' file.


• Confirmation -> Play
  
  Click on 'next' or 'play' button on player to view various position of ligand in docking. These numbers tell ranking of ligand docking confirmation.

Go Back

TOP