Abstract
The reaction of enol(ate)s with electrophiles is used extensively in organic synthesis for stereoselective C–C bond formation. Protein-based catalysts have had comparatively limited application for the stereoselective formation of C–C bonds of choice via enolate chemistry. We describe protein engineering studies on 5-carboxymethylproline synthases, members of the crotonase superfamily, aimed at enabling stereoselective C–C bond formation leading to N-heterocycles via control of trisubstituted enolate intermediates. Active site substitutions, including at the oxyanion binding site, enable the production of substituted N-heterocycles in high diastereomeric excesses via stereocontrolled enolate formation and reaction. The results reveal the potential of the ubiquitous crotonase superfamily as adaptable catalysts for the control of enolate chemistry.
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Acknowledgements
The authors acknowledge the Biotechnology and Biological Sciences Research Council, the Ministry of Higher Education (Egypt, R.B.H.), CONACyT and FIDERH (Mexico, R.G.C.), Cancer Research UK (A.T.) and the Deutsche Akademie der Naturforscher Leopoldina (Germany, C.D.) for financial support. The ccr/pET3d construct was a kind gift from B.E. Alber at the Ohio State University.
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R.B.H. and D.H. identified and prepared CMPSs. R.G.C. prepared Ccr. R.B.H., D.H. and R.G.C. carried out enzyme assays and analysed the results. R.B.H. purified and ran NMR analysis of the products of enzyme catalysis. R.B.H. and T.D.W.C. analysed the NMR data. A.T. synthesized the (2S,5S,6R)-6-ethylcarbapenam methyl ester. C.D. synthesized protected L-AASA (2). R.B.H. and C.J.S. designed the study and wrote the manuscript. All authors commented on the manuscript.
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Hamed, R., Gomez-Castellanos, J., Thalhammer, A. et al. Stereoselective C–C bond formation catalysed by engineered carboxymethylproline synthases. Nature Chem 3, 365–371 (2011). https://doi.org/10.1038/nchem.1011
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DOI: https://doi.org/10.1038/nchem.1011
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