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

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 H₂O₂ 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.