Regio- and Stereoselective Steroid Hydroxylation at C7 by Cytochrome P450 Monooxygenase Mutants.
cytochromes
directed evolution
enzymes
oxidation
steroids
Journal
Angewandte Chemie (International ed. in English)
ISSN: 1521-3773
Titre abrégé: Angew Chem Int Ed Engl
Pays: Germany
ID NLM: 0370543
Informations de publication
Date de publication:
20 07 2020
20 07 2020
Historique:
received:
01
03
2020
revised:
31
03
2020
pubmed:
4
4
2020
medline:
30
3
2021
entrez:
4
4
2020
Statut:
ppublish
Résumé
Steroidal C7β alcohols and their respective esters have shown significant promise as neuroprotective and anti-inflammatory agents to treat chronic neuronal damage like stroke, brain trauma, and cerebral ischemia. Since C7 is spatially far away from any functional groups that could direct C-H activation, these transformations are not readily accessible using modern synthetic organic techniques. Reported here are P450-BM3 mutants that catalyze the oxidative hydroxylation of six different steroids with pronounced C7 regioselectivities and β stereoselectivities, as well as high activities. These challenging transformations were achieved by a focused mutagenesis strategy and application of a novel technology for protein library construction based on DNA assembly and USER (Uracil-Specific Excision Reagent) cloning. Upscaling reactions enabled the purification of the respective steroidal alcohols in moderate to excellent yields. The high-resolution X-ray structure and molecular dynamics simulations of the best mutant unveil the origin of regio- and stereoselectivity.
Identifiants
pubmed: 32243054
doi: 10.1002/anie.202003139
pmc: PMC7384163
doi:
Substances chimiques
Steroids
0
Cytochrome P-450 Enzyme System
9035-51-2
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
12499-12505Informations de copyright
© 2020 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.
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