NCBB Design Server Documentation

Overview

This application performs design of Noncanonical Backbones (NCBB) to act as an inhibitor for a target protein. NCBB's are non-peptide, non-nucleotide based backbones (also known as peptidomimetics). The NCBBs that are available to run are OOPs, HBSs, and peptoids.

Oligooxopiperazines (OOP) are helical mimetics, which mimic one face of an alpha helix and show promise as protein interaction inhibitors. In this mode, minor rigid body perturbations along with puckering of the oop rings and small phi/psi angle moves are iterated with the design of residues on the oop scaffold.

Hydrogen Bond Surrogates (HBS) scaffolds are stapled helices, which converts a backbone hydrogen bond to a covalent bond, stabilizing the molecule in a helix conformation. HBS's show promise as protein interaction inhibitors. In this mode, minor rigid body perturbations are iterated with the design of residues on the HBS scaffold.

Peptoid scaffolds are N-substituted glycine amino acids where side chains are moved one atom over, from the alpha-carbon to the backbone nitrogen. In this mode, minor rigid body perturbations are iterated with the design of residues on the peptoid scaffold.

Inputs

You need to input

Starting model: The model as a pdb file. The user needs to create a rosetta-formatted pdb file (see structure preparation documentation for tips). Here is an OOP example, a HBS example and a Peptoid example. Additionally, the rigid body conformation of the NCBB scaffold with respect to the target needs to be very close (a few angstroms RMSD) to the anticipated binding mode. This application does not do a full docking procedure so the starting conformation is important to get a potential high affinity design. To create a starting model, we therefore recommend manipulating these sample files in a program such as pymol and "pairfit" the scaffold to produce a starting structure that mimicks hotspot residues at an interface.
Design NCBB residues: Residues on NCBB scaffold to be redesigned by Rosetta. Only integers are allowed and each residue should be separated by a space. The numbers should be the index of the residue on the NCBB scaffold (n-term to c-term) (this will be different than the PDB numbering). So for example, the n-term residue is residue 1. Default of no input results in repacking of sidechains with no design.
Is the second chain cyclic? (peptoid only): The peptoid application can handle cyclic peptoid macrocycles and a special procedure is done to keep the macrocyle in a physically realistic conformation. Note: the sample peptoid pdb given above is a cyclic peptoid, so this box should be checked when using that example pdb.
Number of design cycles [Optional]: The number of design (repack) cycles to be done during a single run. Only integers are allowed. Default is 10 cycles.
Number of perturbations per cycle [Optional]: The number of rigid body perturbation steps done during a cycle. Only integers are allowed. Default is 100 perturbations.

Output Files

Output files can be accessed from the job details page. The Redesigned Model PDB is a link to the final model. The Full set of decoy structures created on this run is a link additional output files. In this directory you will find several files.

decoys: Directory that holds all decoys produced during run.
model_redesign.pdb: Final model also linked from job details page.
score.sc: Score file which holds Rosetta score function information for every decoy generated
logs: Directory that holds log file information during run (not generally informative unless an error occured).
*.condor: Internal job files for server function (not generally informative unless an error occured).

Interpreting Results

The server returns one final model as a pdb file along with the full decoy set. The final model is selected from the set of decoys by filtering the top 5% of scoring models and sorting by binding energy. The user can then download and examine the model. It may be of interest for the user to examine more than just the top model because there is often different sequence designs produced with similar energies to the final design. We therefore recommend analyzing the full decoy set for other potential high affinity designs.

Example

To test the server, the user can use the following input of an OOP mimicking p53 targeting MDM2. The input model can be downloaded here. The following inputs can be used:

Design NCBB residues: 1 2 4 (these correspond to p53 residues Phe19, Trp23 and Leu26 respectively)
Number of design cycles: 10
Number of perturbations per cycle: 100

An example output file can be downloaded here (this is a stocastic algorithm so your results may vary).

Power Users

Our aim to provide reproducible results. Below is the commandline options used by the server for the above example:

	./oop_design.static.linuxgccrelease 
		-database ${Rosetta_Database_location} 
		-include_patches patches/oop_pre.txt patches/oop_post.txt   
		-ex1 -ex2 -packing::extrachi_cutoff 0 
		-out:path:pdb output 
		-s ./input/model.pdb 
		-scorefile output/score.sc 
		-multiple_processes_writing_to_one_directory  
		-nstruct 1000 
		-pert_num 100 
		-design_loop_num 10  
		-oop_design_positions 1 2 4

Once the above command is run, you can use the analyze.py script to filter results. In the same directory as your "score.sc" file, you can run this like:

	python analyze.py

Citations

We'd be grateful if you cite our work in publications that make use of the server:

Coming soon.

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