a valid PDB file in which:
The receptor is the first chain
The peptide is the second chain
No heteroatoms are included
The occupancy column is filled (i.e. no double conformations or 0.00 occupancy)
A reference structure (PDB file) to which RMSD will be calculated for the resulting models
Number of models to be created for the high resolution protocol (0-300)
Number of models to be created including the low resolution pre-optimization protocol (0-300)
Selection of additional scores to be reported
Specified constraints to be enforced during the simulation
Note 1: Only AtomPair constraints are supported at the moment
Note 2: The numbering of constraints is sequential and disregards PDB numbering
Example: For a PDB file in which the first chain is a 50 amino acid long protein and the second chain is a 10 amino acid long peptide, enforcement of a distance constraint of 4A +/- 0.5A between the C-alpha atom of residue 23 in the protein and the C-beta atom of residue 3 of the peptide will be achieved with the following syntax:
AtomPair CA 23 CB 53 HARMONIC 4.00 0.5
For more information about the constraints file format, click here.
A zipped file of the top 10 models predicted for this peptide-protein complex (sorted by Rosetta score)
A score file containing the score and RMSD for each model (as well as additional score terms if specified by user)
Top 10 peptides superimposed at the binding site for appreciation of convergence
Separate figures for each of the top 5 models
A plot of the 200 models created by FlexPepDock, showing Energy (y-axis) vs. RMSD from the reference structure (x-axis) (If no reference structure is provided, the RMSD is calculated relative to the submitted starting structure)
Q. I have submitted my job - how long will it take to complete? The low-resolution protocol cannot handle modified residues so using the advanced options, specify the number of low resolution structures as 0, and specify as many high-resolution structures as you like.
Make sure the phosphorylated residue in your PDB file is named properly (i.e. TYR/SER/THR and not PTR/SEP etc...)
Make sure the modified residue coordinates (as well as the phosphates) are marked as ATOM and not HETATM.
Below you can find an example as to how a phospho-serine should look like:
A. Running time depends on the size of the protein, and on the queue. Once FlexPepDock starts running, the job should be completed within 1-2 hours (for a protein of regular size).
Q. I've placed a peptide randomly far away from the protein receptor and I get weird results all over the place ?!
A. Rosetta FlexPepDock is optimized for 'local' peptide docking, i.e. the initial conformation of the peptide should already be located at the approximate binding site. Benchmarking studies showed that it is effective for the refinement of starting structures that deviate by up to 5.5A from the native structure (sometimes even more). However, the initial peptide placement has to be within the approximate binding site.
Q. It doesn't look like there are 200 points in the results plot of my run ?!
A. In order to focus on the more relevant solutions we only display models that scored up to one standard deviation above the average score of all models.
Q. I can not see the interface clearly in the nice figures you provide ?!
A. The figures are created automatically by a PyMOL script, and therefore they sometimes fail to display the complex from the most informative angle. We encourage the users to download the models and view them in their favorite molecular viewer.
Q. Can I dock phosphorylated peptides ?
A. Of course you can! In order to dock phosphorylated peptides make sure you do the following:
ATOM 5776 N SER B 10 -19.024 43.939 120.740 1.00 0.00
ATOM 5777 CA SER B 10 -20.442 43.615 120.653 1.00 0.00
ATOM 5778 C SER B 10 -20.869 42.699 121.792 1.00 0.00
ATOM 5779 O SER B 10 -20.125 41.804 122.194 1.00 0.00
ATOM 5780 CB SER B 10 -20.750 42.972 119.314 1.00 0.00
ATOM 5781 OG SER B 10 -22.089 42.569 119.219 1.00 0.00
ATOM 5782 P SER B 10 -22.461 41.858 117.817 1.00 0.00
ATOM 5783 O1P SER B 10 -24.008 41.465 117.873 1.00 0.00
ATOM 5784 O2P SER B 10 -21.536 40.564 117.675 1.00 0.00
ATOM 5785 O3P SER B 10 -22.170 42.912 116.653 1.00 0.00
Q. What do all the score terms mean ?
A. Here are the details of the different score terms:
score - Total Rosetta energy for this model
fa_atr - Full atom lennard-jones attractive part of Van Der Waals term
fa_rep - Full atom lennard-jones repulsive part of Van Der Waals term
fa_pair - Statistics based pair term that favors salt bridges
fa_sol - Lazaridis-Karplus solvation energy
fa_dun - Internal energy of sidechain rotamers as derived from Dunbrack's statistics
hbond_sc - Sidechain-sidechain hydrogen bond energy
rama - Ramachandran preferences score
I_sc - Interface energy (The total energy of the complex less the energy of the partners when separated)
pep_sc - The sum of energies contributed only by peptide residues
pep_sc_noref - pep_sc less the fixed reference energy for each amino acid in the peptide
All RMS terms are Root Mean Square Deviation calculated to the provided reference structure over the peptide residues. If no reference structure was provided they are calculated against the submitted initial structure:
rmsBB - RMSD is calculated only for peptide backbone heavy atoms
rmsBB_if - RMSD is calculated only for heavy backbone atoms of peptides residues at the interface.
rmsALL_if - RMSD is calculated for all of the heavy atoms of peptides residues at the interface (including side-chains)
startRMSbb - RMSD of starting structure (submitted structure) from the reference structure
Q. What if I want to include a small molecule at the putative interface with the peptide?
A. At the moment the server can not handle small molecules at the interface and will discard them. Future versions will implement this feature.