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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
make
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Install the xmgrace template file, which is an enhanced template
containing more colors that allow for the plotting of the graphs used
here:
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
"to_GLY.side_chains.final.agr". 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.
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