Discovery and Engineering of an Ene‐reductase for Kinetic Resolution and Cascade Reduction of Hajos‐Parrish/Wieland‐Miescher Ketones

Abstract Ene‐reductase (ERED)‐catalyzed asymmetric reduction of α,β‐unsaturated ketones is an attractive method in organic synthesis. Through the discovery and further structure‐guided protein engineering of BsER, an ERED from Bacillus subtilis , the selective bio‐reduction of the well‐known building blocks Hajos‐Parrish ketone (HPK, 1 a ) and Wieland‐Miescher ketone (WMK, 1 b ) was achieved in this study. The optically pure ( R ) ‐ HPK (( R )‐ 1 a ) and ( R ) ‐ WMK (( R )‐ 1 b ) with corresponding >99% and 98% enantiomeric excess ( ee ) values were obtained by kinetic resolution of the racemic substrates. To the best of our knowledge, this is the first example of ERED mediated kinetic resolution of rac ‐HPK and rac ‐WMK. In addition, the reduction products cis ‐7a‐methylhexahydro‐1H‐indene‐1,5(4H)‐dione ( cis ‐ 2 a ) and cis ‐8a‐methylhexahydronaphtha‐ lene‐1,6(2H,5H)‐dione ( cis ‐ 2 b ) with high diastereomeric ratios ( dr ) were also obtained. We also developed four routes of biocatalytic cascade reduction involving BsER and ketoreductases (KREDs). Using optimized ERED‐KRED and KRED‐ERED cascades, ( S ) ‐ HPK (( S )‐ 1 a ) and ( S ) ‐ WMK (( S )‐ 1 b ) could be reduced stepwise, yielding the bioactive products all with >91% purity. This study presents a biocatalytic strategy for the kinetic resolution and cascade reduction of HPK and WMK ( 1 a and 1 b ).