Energy minimization of a protein with NAMD
1 Introduction
2 What's you need
3 NAMD installation
4 Protein download
5 Protein preparation
6 Creation of the input files for NAMD
7 Run the NAMD calculation
8 Analysis of the results
9 Using the atom constraints
9.1 Atom fixing
9.2 Atom constraints
1 Introduction
VEGA ZZ allows to prepare the input files for NAMD in easy way without the
use of complex softwares to generate the topology. In this tutorial is explained
step-by-step how to perform a simple conjugate gradients energy minimization of
the crambin crystallographic structure with and without constraints.
2 What's you need
- VEGA ZZ release 2.0.8 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 Win32-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_Win32-i686 directory in NAMD (X.X is the
NAMD version).
4. Protein download
You can download the crambin (1CRN) structure 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 the PDB Id field and click Download. At
the download end, 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) with
the 1CRN file name. It's strongly recommended the use of the IFF/RIFF
file format because it's able to keep 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 the Molecule type box to enable an extra
check for the atom hybridization, Residue end in the 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 the 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 Creation of the input files for NAMD
NAMD requires several files: the PDB and the PSF file of the molecule, one or
more parameter files and a command file defining the condition of the
calculation.
- Save the molecule in PDB 2.2 format (File
-> Save As...) with the 1CRN.pdb file name. NAMD doesn't need the
PDB connectivity and so you can avoid to save it unchecking Connectivity
in the Options box.
- Create the topological matrix saving the molecule in the X-Plor PSF
format (1CRN.psf file name). Optionally, it's possible to check if all
force field parameters are available, selecting the force field name in the
Force field param. box. The CHARMM22_PROT force field was used in
the atom type attribution and so you must select the same force field. This
operation isn't needed because, the molecule is a standard protein and all
parameters are included. It's useful if you are managing a non-standard
structure in order to fix the missing parameters (for more information,
click here).
- Copy in the directory where are placed the 1CRN.pdb and 1CRN.psf
files, the par_all22_prot.inp and par_all22_vega.inp parameter
files that are in the ...\VEGA ZZ\Data\Parameters directory. The par_all22_vega.inp is
required because VEGA ZZ generates a topology making
explicit all improper angles.
- With a text editor (e.g. Notepad) create the input file with the following
commands (copy & paste them):
numsteps 10000
minimization on
dielectric 1.0
coordinates 1CRN.pdb
outputname 1CRN
outputEnergies 1000
binaryoutput no
DCDFreq 1000
restartFreq 1000
structure 1CRN.psf
paraTypeCharmm on
parameters par_all22_prot.inp
parameters par_all22_vega.inp
exclude scaled1-4
1-4scaling 1.0
switching on
switchdist 8.0
cutoff 12.0
pairlistdist 13.5
margin 0.0
stepspercycle 20
Save the file with the 1CRN_min.namd name. This file allows to
perform a 10000 steps conjugate gradients minimization, saving the output
(coordinates and restart files) every 1000 iterations. For more information about
the parameters, please consult the
NAMD
User Guide.
7 Run the NAMD calculation
- Open the VEGA console (Start -> VEGA ZZ -> VEGA console).
- Go to inside your working directory with the cd command.
- In the console type:
namd2 1CRN_min.namd
and hit return. If you want to save the output in a file, use this command:
namd2 1CRN_min.namd > 1CRN.out
If you have more than one CPU installed, you can speed-up the
calculation specifying the total number of CPUs:
namd2 +p2 1CRN_min.namd > 1CRN.out
In this case the PC has two CPUs and both are used for the calculation (+p2
option). The dual core (e.g. Athlon X2, Pentium D, Core 2 Duo, etc) and the
Pentium 4 with hyperthreading should use the +p2 option.
8 Analysis of the results
The calculation results are contained mainly in two files: the 1CRN.dcd
(trajectory file) and the 1CRN.out. The first one is a binary
file that can't be opened with 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 the 1CRN.dcd file with 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.
- If you saved the 1CRN.out file, the Energy Graph button will
be active. Clicking it, 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). Please remember that in an energy minimization, the last
conformation in the trajectory is always the lowest one and so it doesn't
exist any difference clicking the Last or the Lowest button.
- Save the best conformation (File -> Save As)
in IFF format (1CRN_min.iff).
In the same way, you can save any conformation in the trajectory selecting it
using the horizontal slider or the Frame number field and choosing File
-> Save As in the menu bar.
9. 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.
9.1 Atom fixing
- Start VEGA ZZ and open the 1CRN.iff file.
- Open the Constraints options
window (Edit -> Coordinates -> Constraints).
- Select Fix in the Mode box and Protein backbone in
the Selection box. Finally, click the Apply button and close the
window. The fixed atoms (the backbone) will be colored in blue and the free
atoms in green.
- Save the molecule (File -> Save As) in
PDB 2.2 format, checking Constraints in the Options box and
typing 1CRN_fix.pdb as file name. The atom fixing information is stored
in the B column of the PDB file.
- Repeat the energy minimization with the following command file:
numsteps 10000
minimization on
dielectric 1.0
coordinates 1CRN_fix.pdb
outputname 1CRN_fix
outputEnergies 1000
binaryoutput no
DCDFreq 1000
restartFreq 1000
structure 1CRN.psf
paraTypeCharmm on
parameters par_all22_prot.inp
parameters par_all22_vega.inp
exclude scaled1-4
1-4scaling 1.0
switching on
switchdist 8.0
cutoff 12.0
pairlistdist 13.5
margin 0.0
stepspercycle 20
fixedAtoms on
fixedAtomsCol B
In red are indicated the differences with the
standard minimization file. Please note that the PSF file is the same of the
previous minimization because the molecule isn't changed.
Save the file with 1CRN_fix_min.namd file name.
- Start the minimization, typing in the console:
namd2 1CRN_fix_min.namd > 1CRN_fix.out
- At the end of the minimization, open the trajectory file. Please remember
that you can't open the trajectory file directly, if it doesn't exist the
correspondent molecule file (e.g. 1CRN_fix.iff). As first step, you
must open the molecule and thus select Calculate -> Analysis in the
menu bar. In the Trajectory analysis window, click the open button and
select the DCD file.
- Moving the horizontal slider, you can see that the backbone is kept fixed.
- Select and save the lowest energy conformation as 1CRN_fix_min.iff.
9.2 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.
- Save the molecule (File -> Save As) in
PDB 2.2 format, checking Constraints in the Options box and
typing 1CRN_const.pdb as file name.
- Repeat the energy minimization with the following command file:
numsteps 10000
minimization on
dielectric 1.0
coordinates 1CRN_const.pdb
outputname 1CRN_const
outputEnergies 1000
binaryoutput no
DCDFreq 1000
restartFreq 1000
structure 1CRN.psf
paraTypeCharmm on
parameters par_all22_prot.inp
parameters par_all22_vega.inp
exclude scaled1-4
1-4scaling 1.0
switching on
switchdist 8.0
cutoff 12.0
pairlistdist 13.5
margin 0.0
stepspercycle 20
constraints on
consref 1CRN_const.pdb
conskfile 1CRN_const.pdb
conskcol B
In red are indicated the differences with the
standard minimization file. Save the file with 1CRN_const_min.namd
name.
- Start the minimization, typing in the console:
namd2 1CRN_const_min.namd > 1CRN_const.out
- At the end of the minimization, open the trajectory file, select and save
the lowest energy conformation as 1CRN_const_min.iff.