The scripts and input files that accompany this demo can be found in the demos/public directory of the Rosetta weekly releases.

KEYWORDS: DESIGN INTERFACES

This demo will walk through the steps of designing a protein-protein interface. The goal of this protocol is to predict mutations on Raf that will enable it to bind Rac.

Demo files

README.md
starting_files/
  -- 1c1y.pdb.gz
  -- 2ov2.pdb.gz
rosetta_inputs/
  -- design_script.xml (dock/design script)
  -- raf-rac.pdb (raf-rac starting interface)
scripts/
  -- score_vs_rmsd.R (creates score vs rmsd plot)
output_files/
  -- directory to store output files

Running the Demo

  1. Create model of Raf-Rac interaction.

    • Uncompress pdb files: 
      gunzip 1c1y.pdb.gz
gunzip 2ov2.pdb.gz
    • Open 1c1y and 2ov2 with Pymol.
    • Superimpose chain A of 1c1y with chain A of 2ov2. Pymol command: 
      super 2ov2 and chain A, 1c1y and chain A
    • Select modeled Raf-Rac complex. Pymol command: 
      select raf-rac, (1c1y and chain B)+(2ov2 and chain A)
    • Save Molecule raf-rac. In pymol: 
      File->Save Molecule->raf-rac.pdb

  2. Run dock/design protocol using rosetta_scripts.

    • Change working directory to the output directory. 
      $> cd output_files
    • Run design_script.xml using rosetta_scripts executable. 
      $> $ROSETTA3/bin/rosetta_scripts.macosgccrelease -s ../rosetta_inputs/raf-rac.pdb -parser:protocol ../rosetta_inputs/design_script.xml -in:file:native ../rosetta_inputs/raf-rac.pdb -ex1 -ex2 -ignore_unrecognized_res -nstruct 1 -overwrite
    • Options:
      • -ex1 -ex2: expand rotamer library for chi1 and chi2 angles used in repacking/design
      • -ignore_unrecognized_res: ignores HETATM lines in input PDB file
      • -nstruct 1: specifies how many times the protocol is run (one decoy is output for each run)
      • -overwrite: overwrites decoys from previous runs

    Each run should take about 90 seconds to complete. For production runs, nstruct should be set to 1000 or greater. This protocol returns decoys named raf-rac_####.pdb and a file named score.sc that contains scores for each decoy.

  3. Postprocessing output from dock/design run.

    • To see the sequence changes in chain B due to design grep out chain B: grep " B " raf-rac.pdb > chainBin.pdb then use rosetta_tools/protein_tools/scripts/SequenceProfile.py to diff the sequences

    • Run R script to generate Interface Score vs. Interface RMSD plot. R is easy to install from here as a pkg that self installs (do "which R" to check install first). The above link is fro MacOSX, but R is available for many other operating systems as well.

        CMD BATCH scripts/score_vs_rmsd.R

      Output: score_vs_rmsd.pdf