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A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
Characterization of a neral reductase from Serratia marcescens: Structure-guided engineering and construction of an
Ruishi Xie1, Long Wang1, Shengfang Wu2
1The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, China.
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.
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