Engineering 2‐Deoxy‐D‐ribose‐5‐phosphate Aldolase for <i>anti</i> ‐ and <i>syn</i> ‐Selective Epoxidations of <i>α</i> , <i>β</i> ‐Unsaturated Aldehydes

Abstract The enzyme 2‐deoxy‐D‐ribose‐5‐phosphate aldolase (DERA) naturally catalyzes the reversible aldol addition between acetaldehyde and D‐glyceraldehyde‐3‐phosphate to yield 2‐deoxy‐D‐ribose‐5‐phosphate. Herein we describe the redesign of DERA into a proficient non‐natural peroxygenase that promotes the asymmetric epoxidation of various α , β ‐unsaturated aldehydes. This repurposed aldolase, named DERA‐EP, is able to utilize H 2 O 2 to accomplish both anti ‐ and syn ‐selective epoxidations of various α , β ‐unsaturated aldehydes to give the corresponding epoxides with moderate to high diastereoselectivity (diastereomeric ratio up to 99 : 1) and excellent enantioselectivity (enantiomeric ratio up to 99 : 1). Crystallographic analysis of DERA‐EP in a substrate‐free and substrate‐bound state provides a structural context for the evolved activity, a clear explanation for the high enantioselectivity, and compelling evidence for catalysis via enzyme‐bound iminium ion intermediates. The unprecedented anti ‐selectivity of DERA‐EP with multiple α , β ‐unsaturated aldehydes is complementary to the syn ‐selectivity of previously reported enzyme‐, metal‐ and organo‐catalysts, making DERA‐EP an attractive new asset to the toolbox of epoxidation catalysts.