Active site diversification of P450cam with indole generates catalysts for benzylic oxidation reactions
Paul P Kelly1, Anja Eichler1, Susanne Herter1
1School of Chemistry & Manchester Institute of Biotechnology, The University of Manchester, 131 Princess Street, M1 7DN, Manchester, United Kingdom.
Abstract:
Cytochrome P450 monooxygenases are useful biocatalysts for C-H activation, and there is a need to expand the range of these enzymes beyond what is naturally available. A panel of 93 variants of active self-sufficient P450cam[Tyr96Phe]-RhFRed fusion enzymes with a broad diversity in active site amino acids was developed by screening a large mutant library of 16,500 clones using a simple, highly sensitive colony-based colorimetric screen against indole. These mutants showed distinct fingerprints of activity not only when screened in oxidations of substituted indoles but also for unrelated oxidations such as benzylic hydroxylations.
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