Characterization of a neral reductase from Serratia marcescens: Structure-guided engineering and construction of an efficient whole-cell cascade for (S)-citronellol synthesis.

Abstract

In this study, a previously uncharacterized neral reductase (an ene-reductase), SmNR (Serratia marcescens Neral Reductase), belonging to the Old Yellow Enzyme family, was identified and functionally characterized from Serratia marcescens HBQA7. SmNR efficiently catalyzes the asymmetric reduction of neral to produce (S)-citronellal with an enantiomeric excess (ee) of >99%. SmNR exhibits a preference for NADPH as a coenzyme, and its specific activities toward the two substrates, neral and geranial, are 2.96 U/mg and 0.74 U/mg, respectively. Through semi-rational design, the S272D mutant with improved catalytic performance was obtained. The Km and Vmax of the S272D mutant are 0.08 mM and 4.03 U/mg, respectively, with a catalytic efficiency kcat/Km reaching 33.6 mM-1·s-1. A three-enzyme cascade whole-cell catalytic system, comprising neral reductase, alcohol dehydrogenase, and glucose dehydrogenase, was constructed to achieve efficient conversion of neral to (S)-citronellol, with a (S)-citronellol yield of 87.7% within 8 h.