Rosetta 3.4
Documentation for AnchoredDesign application
Author:
Steven Lewis smlewi@gmail.com

Metadata

Code and documentation by Steven Lewis smlewi@gmail.com. This document was last updated 6/24/11 by Steven Lewis. The PI was Brian Kuhlman, bkuhlman@email.unc.edu.

Examples

The code is at rosetta/rosetta_source/src/apps/public/interface_design/anchored_design/AnchoredDesign.cc; there's an integration test at rosetta/rosetta_tests/integration/tests/AnchoredDesign/. There is a more extensive demo with more documentation at rosetta/rosetta_demos/AnchoredDesign, or in the demo section of the release.

References

Purpose and Algorithm

AnchoredDesign is the main protocol discussed here. This protocol is meant to design interfaces between known target structures and new binding partners, using an "anchor" consisting of residues grafted from a known binding partner of the target onto the new scaffold. The idea is that this will reduce the conformational space we need to search and preclude the docking problem, while still leaving freedom to design the interface as necessary. The anchor should be grafted into a surface loop of the scaffold (see Documentation for AnchoredPDBCreator application). Loop remodeling of the anchor loop will move the scaffold with respect to the target, exposing different parts of their surfaces to one another. Loop remodeling of other (unanchored) surface loops is also implemented. This lets us design a new, flexible-backbone interface between new binding partners.

Input Files

See rosetta/rosetta_tests/integration/tests/AnchoredDesign/ for example usage.

AnchoredDesign requires as inputs a starting structure, an anchor specification, and a loops specification (Documentation for the loop modeling application). The starting structure needs to have the anchor grafted into the scaffold (see Documentation for AnchoredPDBCreator application and its documentation), and the anchor needs to be docked properly relative to the target. The anchor/target interaction WILL NOT CHANGE, since it was drawn from a crystal structure. The relationship between the scaffold and rest of the system is treated flexibly; the scaffold does need to be connected to the anchor but it's fine if the scaffold crashes horribly into the target (that will be worked out by the protocol).

Note that the AnchoredDesign protocol respects the start, end, cut, and extended fields of the loop file. It ignores the skip_rate field.

The anchor file specification is a one-line file with three whitespace-delimited values: the chain letter, start, and end residue of the anchor. Like so:

B 442 445

It's going to assume PDBInfo exists, so if you have silent files try numbering from 1.

Options

AnchoredDesign has its own namespace of options, and also supports general Rosetta options (packing, etc.)

AnchoredDesign options

These four boolean options allow deactivation of KIC or CCD loop closure for perturb and refine stages:

AnchoredDesign has a sub-namespace for filtering:

These options activate vicinity sampling in kinematic loop closure. (The default is to use random samples from the Ramachandran distribution for the non-pivot torsions in the loop; these options instead perturb the existing loop slightly)

General options: All packing namespace options loaded by the PackerTask are respected. jd2 namespace options are respected. Anything very low-level, like the database paths, is respected.

PoseMetricCalculator flags include:

Tips

fluorophore modeling

This section describes changes for the fluorophore modeling experiments (

Anchoring via constraint

The code is capable of holding the anchor in place via constraints, rather than rigid locking through the fold tree. It will maintain a rigid anchor in the centroid perturb phase no matter what (I don't trust the centroid scorefunction that much), but it will allow internal backbone movement of the anchor, as well as rigid body movement, in full atom mode. I suggest using tight strong constraints to keep your anchor in place. Use these flags:

Expected Outputs

AnchoredDesign should produce nstruct worth of result structures. If you used the default JobOutputter, you'll get PDBs with embedded score information and a scorefile summarizing most of the score information.

Post Processing

There are three classes of output tagged to the end of the PDBs and/or in the scorefile:

The scorefunction tells you what Rosetta's standard scorefunctions think is best, and is useful for the first sorting of structures. Generally, only structures in the top few percent by total_score should be further analyzed. Even then, the other scorefunction terms (listed at the end of the PDB and in the score file) should be examined to throw out models that have particularly bad scores in any area - a model that is overall pretty good, but has a bad clash (fa_rep) for one particular residue, may be worth throwing out.

Next, you use the LoopAnalyzerMover output to filter for bad loop closures (which Rosetta's scorefunction detects insufficiently). LoopAnalyzerMover tags this output to the end of the PDB. The second line is long column titles, and the third is short versions to make visualization easier. Each row represents one residue. Totals for all loops for some terms are collected at the bottom.

LoopAnalyzerMover: unweighted bonded terms and angles (in degrees)
position phi_angle psi_angle omega_angle peptide_bond_C-N_distance rama_score omega_score dunbrack_score peptide_bond_score chainbreak_score
 pos phi_ang psi_ang omega_ang pbnd_dst    rama  omega_sc dbrack pbnd_sc   cbreak
  17  -106.8   175.8     178.2    1.322   0.998    0.0342   7.01   -2.68   0.0182
  18  -82.33   64.67    -178.5    1.329   0.211    0.0217   3.11   -3.42   0.0203
  19  -83.63   149.4     177.2    1.329   -1.07    0.0795      0   -3.43    0.584
  20  -75.25   171.1    -178.7    1.329  -0.264    0.0161  0.348   -3.43   0.0151
  21  -58.53  -42.95     174.6    1.329   -0.58     0.294      0   -3.43      2.7
  22  -76.02   159.9    -179.8    1.326  -0.811  0.000404   0.97   -3.45   0.0424
  23  -72.63   130.1     179.4    1.325   -1.29   0.00372   0.24   -3.46   0.0281
  24  -94.91   116.5     179.8    1.323   -1.21   0.00028  0.721   -3.45   0.0694
  25  -65.42   150.7     179.4    1.335   -1.58     0.004      0   -3.32     1.38
  26  -64.68   147.9     179.1    1.323   -1.45    0.0079   1.61   -3.32    0.211
  27  -56.44  -66.68      -180    1.329    1.34  8.08e-30   7.87   -3.43 2.37e-05
  28  -124.4  -56.48     177.6    1.329    2.08    0.0568  0.608   -3.43   0.0533
  29  -124.1   28.78    -177.7    1.264   0.341    0.0542   2.39    2.65     2.07
  30   81.57  -134.3    -176.4    1.329      20     0.126   5.06    2.65    0.128
  31  -112.9   147.2     172.7    1.318  -0.744     0.538  0.534   -3.35     1.38
total_rama 15.9674
total_omega 1.23676
total_peptide_bond -38.3223
total_chainbreak 8.70689
total rama+omega+peptide bond+chainbreak -12.4113

LAM_total -12.4113

In this particular example, position 29 is clearly problematic: the peptide bond distance is too short, as reported by the pbnd_dst, pbnd_sc, and cbreak columns. You can also see that position 30 has an awful ramachandran score. Good structures will have no fields out of range of the lower scores in this example.

Finally, InterfaceAnalyzer puts results at the end of the PDB file as well; read about it here: Documentation for InterfaceAnalyzer application. Again, throw out models with poor interfaces as determined by InterfaceAnalyzer.

Changes since last release

Rosetta 3.3 is the first release.

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