Institute of Chemistry - University of Campinas

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How to use the scripts:

1. Prepare a structure that can actually be simulated with NAMD

Every ATD simulation starts from a pre-equilibrated structure. Therefore, you must have a system setup which can be run with NAMD, as if you were about to run a production simulation. This is a prerequisite for using these scripts, and how to do that is described in NAMD tutorials and manuals. Anyway, you probably should be familiar with molecular dynamics simulations before trying to use novel techniques.

2. Setup the ATD scripts:

You must have TCL, gfortran and xmgrace installed in your system. And we suppose you are running some linux flavor.
tar -xzvf atd_scripts.VERSION.tar.gz
cd atd_scripts
Install the xmgrace template file, which is an enhanced template containing more colors that allow for the plotting of the graphs used here:
make grace
Note: This will override your ~/.grace/templates/Default.agr file, and copy it to Default.agr.BAK. Generally that is fine, but if you have changed your Default.agr for some reason, backup it.

3. Modify the input files:

The "atd_scripts/input_example" directory contains three files, which exemplify how the ATD scripts are to be run:

trbeta_equilibrated.pdb: This is the structure file, which is a PDB file which contains an equilibrated molecular system, probably extracted from a previous equilibrium simulation.

trbeta.psfgen: This is the psfgen script that must be modified so that the ATD scripts are able to build all the simulated systems. It must generate, at the end, adequate PSF and PDB files for your system, using the PDB file provided above and the topology files which must be provided. This file must be modified according to your system, except where the example file clearly indicates you must not.

atd_script.inp: This is the main file that will be used as input for all scripts provided. You must modify it by providing the correct path for the psfgen file above, the parameter files, and psfgen and namd2 executables. Also, you must modify the path to the standard NAMD input file, described below.

NAMD_standard_ATD_input.namd: This is the NAMD input file for running each ATD simulation. Generally, you will not need to modify it, but of course you may want to change simulation parameters. In that case, you can do modifications, always preserving the default input and output names (in capital letters), which are used by the scripts.

4. Run ATD simulations

Go to the directory were your "atd_scripts.inp" file is located. You will run everything from there.

Then, you have to run the scripts by using the following steps:

4.1. Setup input files
atd_prepare.tcl atd_scripts.inp [native/GLY/ALA...]

This script will create the input files for running ATD simulations for every residue of your protein, either in native state, or mutated (one by one) to any other residue, for example GLY or ALA. For example, in order to prepare input for GLY mutants, run:

atd_prepare.tcl atd_scripts.inp GLY

4.2. Running simulations
atd_run.tcl atd_scripts.inp GLY

The input files for the simulations were created by atd_prepare.tcl. Now you actually run the ATD simulations using atd_run.tcl, using the same syntax. Actually, after this is script is finished, the ATD simulations are done. Next, there are some scripts for data analysis.

4.3. Computing temperatures
atd_temperatures.tcl atd_script.inp GLY

Using the same input syntax, this script will compute for every simulation the temperature of every other residue along the simulation. It creates on the directory of every simulation a "temperature.dat" file, containing this information, which is used for the subsequent analyzes.

4.4. Create the global thermal diffusion plot
atd_total.tcl atd_script.inp [native/GLY/ALA...]

This script will read the output of the previous temperature run and plot the final and average temperatures of the protein as a function of the heating of each residue. Also, it writes a file containing the final temperature of the protein ordered from higher to lower according the residue heated. The three files are output to the output/graphs directory.

4.5. Create maps of thermal diffusion
atd_map.tcl atd_script.inp GLY [temp_min] [temp_max]

This script will create the thermal diffusion maps for the simulations specified (in this case, the GLY mutatns). You can adjust the color scale of the map with the optional [temp_min] and [temp_max] values. The map created is xmgrace file (.agr), which will be located at the "output/graphs" directory.

Note: In order to view color scale correctly with xmgrace, you need to put the "Default.agr" file, provided with the scripts, in the "~/.grace/templates/" directory.

4.6. Compute side-chain contributions
atd_sidechains.tcl atd_script.inp GLY

After you have run both a mutant (GLY or ALA or something else) and the native simulation, you can compute the side chain contributions for thermal diffusion. This script does that. Running it will produce a graph, also for xmgrace, which will be placed also on the "output/graphs" directory, and will be called something like "to_GLY.side_chains_average.agr" or "". The "average" graph contains the side chain contributions to the average temperature of the protein during the ATD simulation, and the "final" graph contains the contribution to the final temperature of the protein at the ATD run (remember that ATD simulations are out of equilibrium, thus the "average" temperature is not necessarily more meaningful than that the final temperature).

4.7. Scripts to monitor and control running simulations:

atd_check.tcl atd_script.inp GLY

This script will check the running status of the simulations, in this case the "GLY" mutant simulations. It will report how many of them are finished, running, or crashed.

atd_clean.tcl atd_script.inp GLY

This script will remove all files created by a previous ATD run for some simulation. Usually you run it if you thing something went very wrong. Use with care.

5. Analysis

The plots created in the "output/graphs" directories contain all the analyzes performed in the reference paper above. Have fun looking at them and trying to find something meaningful, we hope so! Good luck.

6. Time dependence of heat propagation

In order to create a plot of the time-dependence of heat propagation, as the graph in Figure 4C of the JPCL article, the "time_dep" program is provided.

A simulation must be run in which a the temperature coupling is assigned to whatever group you want to heat, and you need to generate a velocity trajectory file using the "velDCDfile" input parameter in your namd2 input file, as, for example:

velDCDfile velocities.dcd

When the simulation is finished, you will run the time_dep program using:

time_dep simulation.psf velocities.dcd PROT 0. 300. > time_dep.dat

Where "simulation.psf" is the simulation PSF file, velocities.dcd is the velocity trajectory file, PROT is the name of the segment for which the map will be built, and 0. and 300. are the minimum and maximum temperatures of the scale. time_dep.dat file will be created and will contain a list of the temperature and color for every residue in every frame.

In order to create the plot, open xmgrace and:

1. Import the time_dep.dat file as "Block Data" with the "XYCOLOR" option, where X is column 1 (the time), Y1 is column 2 (the residue number) and Y2 is column 4 (the color).

2. Remove the line (Line properties; Type:None)

3. Add "Square Symbols". In the "Symbols" tab, remove the Symbol outline (Style:None) and chose Pattern:Solid Black.

4. Adjust the size of the symbols, the scale and the width of the graph in order that the plot looks nice.

The colors are in a scale from 200 (blue) to 226 (red), according to the definitions of the Default.agr file which is installed using the "make grace" command, explained above.
See also:
Author's software page
The TANGO project