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  P450-Catalyzed Regio- and Diastereoselective Steroid Hydroxylation: Efficient Directed Evolution Enabled by Mutability Landscaping

Acevedo-Rocha, C. G., Gamble, C. G., Lonsdale, R., Li, A., Nett, N., Hoebenreich, S., et al. (2018). P450-Catalyzed Regio- and Diastereoselective Steroid Hydroxylation: Efficient Directed Evolution Enabled by Mutability Landscaping. ACS Catalysis, 8(4), 3395-3410. doi:10.1021/acscatal.8b00389.

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Acevedo-Rocha, Carlos G.1, 2, Autor           
Gamble, Charles G.3, Autor
Lonsdale, Richard1, 2, 4, Autor           
Li, Aitao1, 2, 5, Autor           
Nett, Nathalie2, Autor
Hoebenreich, Sabrina2, Autor
Lingnau, Julia B.6, Autor           
Wirtz, Cornelia6, Autor           
Fares, Christophe6, Autor           
Hinrichs, Heike7, Autor           
Deege, Alfred7, Autor           
Mulholland, Adrian J.4, Autor
Nov, Yuval8, Autor
Leys, David3, Autor
McLean, Kirsty J.3, Autor
Munro, Andrew W.3, Autor
Reetz, Manfred T.1, 2, Autor           
Affiliations:
1Research Department Reetz, Max-Planck-Institut für Kohlenforschung, Max Planck Society, ou_1445588              
2Department of Chemistry, Philipps-University, Hans-Meerwein-Strasse 4, 35032 Marburg, Germany, ou_persistent22              
3Manchester Institute of Biotechnology, School of Chemistry, University of Manchester, Manchester M1 7DN, U.K., ou_persistent22              
4Centre for Computational Chemistry, School of Chemistry, University of Bristol, Cantock’s Close, Bristol BS8 1TS, U.K., ou_persistent22              
5Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, 368 Youyi Road, Wuchang Wuhan 430062, China, ou_persistent22              
6Service Department Farès (NMR), Max-Planck-Institut für Kohlenforschung, Max Planck Society, ou_1445623              
7Service Department Schulze (GC, HPLC), Max-Planck-Institut für Kohlenforschung, Max Planck Society, ou_persistent22              
8Department of Statistics, University of Haifa, Haifa 31905, Israel, ou_persistent22              

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Schlagwörter: directed evolution; cytochrome P450 monooxygenase; regioselectivity; stereoselectivity; mutability landscapes; iterative saturation mutagenesis; steroids; C-H activiation
 Zusammenfassung: Cytochrome P450 monooxygenases play a crucial role in the biosynthesis of many natural products and in the human metabolism of numerous pharmaceuticals. This has inspired synthetic organic and medicinal chemists to exploit them as catalysts in regio- and stereoselective CH-activating oxidation of structurally simple and complex organic compounds such as steroids. However, levels of regio- and stereoselectivity as well as activity are not routinely high enough for real applications. Protein engineering using rational design or directed evolution has helped in many respects, but simultaneous engineering of multiple catalytic traits such as activity, regioselectivity, and stereoselectivity, while overcoming trade-offs and diminishing returns, remains a challenge. Here we show that the exploitation of information derived from mutability landscapes and molecular dynamics simulations for rationally designing iterative saturation mutagenesis constitutes a viable directed evolution strategy. This combined approach is illustrated by the evolution of P450BM3 mutants which enable nearly perfect regio- and diastereoselective hydroxylation of five different steroids specifically at the C16-position with unusually high activity, while avoiding activity–selectivity trade-offs as well as keeping the screening effort relatively low. The C16 alcohols are of practical interest as components of biologically active glucocorticoids.

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Sprache(n): eng - English
 Datum: 2018-01-292018-03-082018-04-06
 Publikationsstatus: Online veröffentlicht
 Seiten: 16
 Ort, Verlag, Ausgabe: -
 Inhaltsverzeichnis: -
 Art der Begutachtung: Expertenbegutachtung
 Identifikatoren: DOI: 10.1021/acscatal.8b00389
 Art des Abschluß: -

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Titel: ACS Catalysis
  Kurztitel : ACS Catal.
Genre der Quelle: Zeitschrift
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Affiliations:
Ort, Verlag, Ausgabe: Washington, DC : ACS
Seiten: - Band / Heft: 8 (4) Artikelnummer: - Start- / Endseite: 3395 - 3410 Identifikator: Anderer: 2155-5435
CoNE: https://pure.mpg.de/cone/journals/resource/2155-5435