Computational design of enone-binding proteins with catalytic activity for the Morita-Baylis-Hillman reaction

TitleComputational design of enone-binding proteins with catalytic activity for the Morita-Baylis-Hillman reaction
Publication TypeJournal Article
Year of Publication2013
AuthorsBjelic, S., Nivón L. G., Çelebi-Ölçüm N., Kiss G., Rosewall C. F., Lovick H. M., Ingalls E. L., Gallaher J. L., Seetharaman J., Lew S., Montelione G. T., Hunt J. F., Michael F. E., Houk K. N., & Baker D.
JournalACS chemical biology
Volume8
Issue4
Pagination749-57
Date Published2013 Apr 19
ISSN1554-8937
KeywordsCollaborative Publication
Abstract

The Morita-Baylis-Hillman reaction forms a carbon-carbon bond between the α-carbon of a conjugated carbonyl compound and a carbon electrophile. The reaction mechanism involves Michael addition of a nucleophile catalyst at the carbonyl β-carbon, followed by bond formation with the electrophile and catalyst disassociation to release the product. We used Rosetta to design 48 proteins containing active sites predicted to carry out this mechanism, of which two show catalytic activity by mass spectrometry (MS). Substrate labeling measured by MS and site-directed mutagenesis experiments show that the designed active-site residues are responsible for activity, although rate acceleration over background is modest. To characterize the designed proteins, we developed a fluorescence-based screen for intermediate formation in cell lysates, carried out microsecond molecular dynamics simulations, and solved X-ray crystal structures. These data indicate a partially formed active site and suggest several clear avenues for designing more active catalysts.

DOI10.1021/cb3006227
Custom1

http://www.ncbi.nlm.nih.gov/pubmed/23330600?dopt=Abstract

Alternate JournalACS Chem. Biol.
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