98
with the –dssp_name switch. Finally, GROMACS versions
prior to 5.0 are not supported.
11. By default, simulations.sh prepares, equilibrates, and runs all
simulations it is asked to without taking a break. However, if it
is terminated, it can restart from the beginning of the last step
it finished successfully, so it may be used (with some modification) as a resubmit script for systems like PBS. If preferred, the
-o switch can be supplied to the script, which will perform the
setup steps and generate individual run folders, but will not
perform the full-length production runs. The full-length simulations can then be run on whatever hardware is appropriate by
simply running gmx grompp on the provided .MDP files and
starting structures.
The script first prepares the MARTINI coarse-grained
force field topology, and converts the provided .PDB file to a
coarse- grained model with the script Martinize. This includes
construction of an elastic network around both protein
domains, which keep their conformations constant and allow
sampling of linker collapse. It then energy minimizes and solvates the model, including addition of neutralizing ions, antifreeze MARTINI water, and experimental salt concentration if
desired. It performs a second energy minimization, and then
equilibrates the system with thermostat and barostat in several
rounds of progressively weaker backbone-restrained MD. Force
constants of 1000, 500, 100, 50, and 10 are used by the script.
The production run involves 30 replicate 200 ns simulations.
New velocities are generated for each of these runs in a final
unrestrained equilibration step. These simulations take approximately 90 min each for a 600 residue fusion model running
on dual Tesla K40s with 32 cores. Finally, it outputs the last
500 ns of each simulation, sampled in 1 ns intervals, as a PDB
file to the results directory.
For instance, if step 2 was performed in directories called
~/dyes/open and ~/dyes/closed, and the script is being run
in ~/dyes, per the example in Note 8, a typical call for a fusion
model with the binding protein occupying residues 242–586
and the fluorophore at residue 63 might be:
python process-data.py open/results/*.
pdb closed/results/*.pdb -f 63 -r 242-586
12. While cysteine mutagenesis is a functional method of chemically labeling a protein with a synthetic dye, alternative chemistries are possible through the use of unnatural amino acid
incorporation. This can allow for biorthogonal labeling of
proteins in vivo, and in principle, it can allow for the sensor to
be genetically encoded in an organism capable of utilizing the
required unnatural amino acid.
Joshua A. Mitchell et al.
with the –dssp_name switch. Finally, GROMACS versions
prior to 5.0 are not supported.
11. By default, simulations.sh prepares, equilibrates, and runs all
simulations it is asked to without taking a break. However, if it
is terminated, it can restart from the beginning of the last step
it finished successfully, so it may be used (with some modification) as a resubmit script for systems like PBS. If preferred, the
-o switch can be supplied to the script, which will perform the
setup steps and generate individual run folders, but will not
perform the full-length production runs. The full-length simulations can then be run on whatever hardware is appropriate by
simply running gmx grompp on the provided .MDP files and
starting structures.
The script first prepares the MARTINI coarse-grained
force field topology, and converts the provided .PDB file to a
coarse- grained model with the script Martinize. This includes
construction of an elastic network around both protein
domains, which keep their conformations constant and allow
sampling of linker collapse. It then energy minimizes and solvates the model, including addition of neutralizing ions, antifreeze MARTINI water, and experimental salt concentration if
desired. It performs a second energy minimization, and then
equilibrates the system with thermostat and barostat in several
rounds of progressively weaker backbone-restrained MD. Force
constants of 1000, 500, 100, 50, and 10 are used by the script.
The production run involves 30 replicate 200 ns simulations.
New velocities are generated for each of these runs in a final
unrestrained equilibration step. These simulations take approximately 90 min each for a 600 residue fusion model running
on dual Tesla K40s with 32 cores. Finally, it outputs the last
500 ns of each simulation, sampled in 1 ns intervals, as a PDB
file to the results directory.
For instance, if step 2 was performed in directories called
~/dyes/open and ~/dyes/closed, and the script is being run
in ~/dyes, per the example in Note 8, a typical call for a fusion
model with the binding protein occupying residues 242–586
and the fluorophore at residue 63 might be:
python process-data.py open/results/*.
pdb closed/results/*.pdb -f 63 -r 242-586
12. While cysteine mutagenesis is a functional method of chemically labeling a protein with a synthetic dye, alternative chemistries are possible through the use of unnatural amino acid
incorporation. This can allow for biorthogonal labeling of
proteins in vivo, and in principle, it can allow for the sensor to
be genetically encoded in an organism capable of utilizing the
required unnatural amino acid.
Joshua A. Mitchell et al.