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Published August 14, 2015 | public
Journal Article

Using molecular dynamics simulations as an aid in the prediction of domain swapping of computationally designed protein variants


In standard implementations of computational protein design, a positive-design approach is used to predict sequences that will be stable on a given backbone structure. Possible competing states are typically not considered, primarily because appropriate structural models are not available. One potential competing state, the domain-swapped dimer, is especially compelling because it is often nearly identical to its monomeric counterpart, differing by just a few mutations in a hinge region. Molecular dynamics (MD) simulations provide a computational method to sample different conformational states of a structure. Here, we tested whether MD simulations could be used as a post-design screening tool to identify sequence mutations leading to domain-swapped dimers. We hypothesized that a successful computationally-designed sequence would have backbone structure and dynamics characteristics similar to that of the input structure, and that in contrast, domain-swapped dimers would exhibit increased backbone flexibility and/or altered structure in the hinge-loop region to accommodate the large conformational change required for domain swapping. While attempting to engineer a homodimer from a 51 amino acid fragment of the monomeric protein engrailed homeodomain (ENH), we had instead generated a domain-swapped dimer (ENH_DsD). MD simulations on these proteins showed increased MD simulation derived B factors in the hinge loop of the ENH_DsD domain-swapped dimer relative to monomeric ENH. Two point mutants of ENH_DsD designed to recover the monomeric fold were then tested with an MD simulation protocol. The MD simulations suggested that one of these mutants would adopt the target monomeric structure, which was subsequently confirmed by X-ray crystallography.

Additional Information

© 2015 Elsevier Ltd. Received 11 March 2015, Revised 11 June 2015, Accepted 16 June 2015, Available online 21 June 2015. The authors are grateful for the use of beamline 12-2 at the Stanford Synchrotron Radiation Lightsource (SSRL) in Menlo Park, CA, operated by Stanford University and supported by the Department of Energy and the National Institutes of Health. We also thank Jens Kaiser and Pavle Nikolovski at the California Institute of Technology for their advice on crystallography. We thank the Gordon and Betty Moore Foundation for support of the Molecular Observatory at the California Institute of Technology. This work was supported by the Defense Advanced Research Projects Agency Protein Design Processes Program, a National Security Science and Engineering Faculty Fellowship (NSSEFF N00244-09-1-0011, N00244-09-1-0082) and the Gordon and Betty Moore Foundation through Grant GBMF2809 to the Caltech Programmable Molecular Technology Initiative. We are grateful to Marie Ary for assistance with the manuscript.

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