Energy minimization of a protein with NAMD

1. Introduction
2. What's you need
3. NAMD installation
4. Protein download
5. Protein preparation
6. NAMD minimization
7. Analysis of the results
9. Using the atom constraints
    9.1 Atom fixing
    9.2 Atom constraints

1 Introduction

VEGA ZZ allows to run NAMD calculations in easy way without the use of external software. In this tutorial, it will explained step-by-step how to perform a simple conjugate gradients energy minimization of the crambin crystallographic structure with constraints.

2 What you need

• VEGA ZZ release 3.0.0 or greater (click here for the software setup).
• NAMD for Windows (click here to download it).
• Test protein. In this tutorial will be used the crystallographic structure of the crambin (1CRN) available at Protein Data Bank (PDB).

3 NAMD installation

• Download the WinYY-i686 package from the Theoretical and Computational Biophysics Group web site.
• Unzip the package in the VEGA ZZ installation directory (usually C:\Program Files\VEGA ZZ).
• Rename the NAMD_X.X_WinYY-i686 directory in NAMD (X.X is the NAMD version).

4 Protein download

You can download the crambin (1CRN) structure by using the PDB Web interface or the tool integrated in VEGA ZZ:

• Start VEGA ZZ and select the File PDB download menu item.
• Put 1CRN in PDB Id field and click Download. At the end of the download, the protein structure will be shown in the workspace (for more information, click here).
• Normalize the coordinates in order to translate the protein at the origin of the Cartesian axis (Edit Coordinates Normalize).
• Save the molecule (File Save As) as 1CRN. It's strongly recommended the use of IFF/RIFF file format because it can include  the maximum number of information (e.g. atom types, charges, bond orders, etc).

5 Protein preparation

• Add the hydrogens (Edit Add Hydrogens), selecting Protein in Molecule type box to enable an extra check for the atom hybridization, Residue end in Position of hydrogens box and checking Use IUPAC atom nomenclature. Finally, click Add to place the hydrogens. Two warning messages will be shown in the console to inform you that two atoms have an unusual geometry and the extra check has corrected the atom type.
• Fix the atom types and the charges (Calculate Charge & Pot.), checking Force field and Charges and selecting CHARMM22_PROT and CHARMM22_CHAR. Click Fix button. The total charge is 0. It's possible to assign the Gasteiger-Marsili charges selecting Gasteiger in the Charges box. This method could be required if the molecule contains non standard residues that aren't included in the CHARMM 22 charges database.
• Save the molecule in IFF format overwriting the previous one.

6 NAMD minimization

All input files required by NAMD are automatically generated by VEGA ZZ.

• Open NAMD  dialog window (Calculate NAMD).
• Go to Other tab, double-click Min - All free (generic) in Presettings box. The NAMD configuration loaded message will be shown in VEGA ZZ console to confirm the operation. In this way, instead to set manually each parameter for a energy minimization, the generic settings are loaded from a generic template file.
• Go to Basic tab and set Number of timesteps to 10000 that in this specific case is the number of minimization steps.
• Go to Input tab and select CHARMM22_PROT as Force field.
• Go to Output tab and uncheck Remove files after minimization.
• In Run mode box, check Run NAMD in interactive mode.
• Finally, click Run to start the minimization.
• If you need to abort the calculation, right-click the workspace and select Stop calculation.
• At the end of the calculation, the structure corresponding to the last minimization step is kept in the workspace. Usually, if the number of minimization steps is large enough, it's also the lowest energy conformation.
• You can save the best conformation (File Save As) in IFF format (1CRN_min.iff).

7 Analysis of the results

The calculation results are placed in the same directory in which you saved the crambin structure and the most interesting files are two: 1CRN.dcd (trajectory file) and 1CRN.out. The first one is a binary file that can't be opened by a text editor. It contains the atom coordinates of each saved frame (10 frames, because one frame every 1000 was saved). The second one is a text files containing the output messages generated by NAMD and the energy information.

• Open 1CRN.dcd file in VEGA ZZ (File Open). To open a DCD trajectory, a molecule file is required (e.g. in PDB or IFF format) with the same name. You shouldn't have any problem because you have the 1CRN.dcd and the 1CRN.iff file in the same directory.
  The molecule will be shown and the Trajectory analysis dialog will be opened.
• Clicking Energy Graph button, you can see the energy behaviour during the calculation. The energy go down as you should expect by an minimization.
• Clicking the Last or the Lowest button in the Trajectory analysis window, the lowest energy conformation is selected (see the workspace).
• Use the horizontal slider or the Frame number field to show the other conformations.

8. Using the atom constraints

In order to keep the structure more close to the original crystallographic data, a common procedure is to apply atom constraints to the protein backbone. In this way, the side chains can relax themselves keeping the secondary structure. NAMD and VEGA ZZ allow to constraint the atoms in two modes: fixing the atoms or applying a force constant to the atoms restraining their movements.

8.1 Atom fixing

• Start VEGA ZZ and open the 1CRN.iff file.
• Open the Constraints options window (Edit Coordinates Constraints).
• Select Fix in Mode box and Protein backbone in Selection box. Finally, click Apply button and close the window. The fixed atoms (the backbone) will be colored in blue and the free atoms in green.
• Repeat the energy minimization as explained in section 6 but instead of Min - All free (generic) preset use Min - Fixed atoms.
• Run the minimization and save the final structure as 1CRN_fix_min.iff.

8.2 Atom constraints

• As above, start VEGA ZZ, open the crambin and show the Constraint options dialog window.
• Select Value in the Mode box, put 20 in the Value field of the Parameters box and choose Protein backbone in the Selection box. Click the Apply button and close the window. If the Value field is 0, the atoms are considered totally free (it means a constraint force constant equal to zero), and increasing that value, the atoms are progressively restrained.
• Repeat the energy minimization as explained in section 6 but instead of Min - All free (generic) preset use Min - Constraints.
• Run the minimization and save the final structure as 1CRN_const_min.iff.