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Enantioselective Epoxidation of Terminal Alkenes to ( R )‐ and ( S )‐Epoxides by Engineered Cytochromes P450 BM‐3
Author(s) -
Kubo Takafumi,
Peters Matthew W.,
Meinhold Peter,
Arnold Frances H.
Publication year - 2006
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/chem.200500584
Subject(s) - bacillus megaterium , enantioselective synthesis , saturated mutagenesis , epoxide , chemistry , cytochrome p450 , stereochemistry , alcohol dehydrogenase , biocatalysis , mutagenesis , catalysis , cofactor , enantiomeric excess , directed evolution , biochemistry , alcohol , enzyme , mutation , biology , bacteria , reaction mechanism , mutant , gene , genetics
Cytochrome P450 BM‐3 from Bacillus megaterium was engineered for enantioselective epoxidation of simple terminal alkenes. Screening saturation mutagenesis libraries, in which mutations were introduced in the active site of an engineered P450, followed by recombination of beneficial mutations generated two P450 BM‐3 variants that convert a range of terminal alkenes to either ( R ) ‐ or ( S ) ‐ epoxide (up to 83 % ee ) with high catalytic turnovers (up to 1370) and high epoxidation selectivities (up to 95 %). A biocatalytic system using E. coli lysates containing P450 variants as the epoxidation catalysts and in vitro NADPH regeneration by the alcohol dehydrogenase from Thermoanaerobium brockii generates each of the epoxide enantiomers, without additional cofactor.