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Enantioselective Reduction of Citral Isomers in NCR Ene Reductase: Analysis of an Active‐Site Mutant Library
Author(s) -
Kress Nico,
Rapp Johanna,
Hauer Bernhard
Publication year - 2017
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201700011
Subject(s) - citral , chemistry , citronellal , selectivity , stereochemistry , active site , docking (animal) , mutant , enzyme , organic chemistry , biochemistry , catalysis , chromatography , medicine , nursing , essential oil , gene
A deeper understanding of the >99 % S ‐selective reduction of both isomers of citral catalyzed by NCR ene reductase was achieved by active‐site mutational studies and docking simulation. Though structurally similar, the E / Z isomers of citral showed a significantly varying selectivity response to introduced mutations. Although it was possible to invert ( E )‐citral reduction enantioselectivity to ee 46 % ( R ) by introducing mutation W66A, for ( Z )‐citral it remained ≥88 % ( S ) for all single‐residue variants. Residue 66 seems to act as a lever for opposite binding modes. This was underlined by a W66A‐based double‐mutant library that enhanced the ( E )‐citral derived enantioselectivity to 63 % ( R ) and significantly lowered the S selectivity for ( Z )‐citral to 44 % ( S ). Formation of ( R )‐citronellal from an ( E / Z )‐citral mixture is a desire in industrial (−)‐menthol synthesis. Our findings pave the way for a rational enzyme engineering solution.