Coimmobilized Dual\nEnzymes in a Continuous Flow Reactor\nfor the Efficient Synthesis of Optically Pure γ/δ-Lactones

Enzyme catalysis is a promising method for producing\nchiral chemicals\nwith high stereoselectivity under mild conditions. However, the traditional\nbatch reaction suffers from low enzyme stability, low cofactor recycling,\nand poor enzyme reusability. Here, we present a continuous-flow method\nusing coimmobilized dual enzymes for the synthesis of chiral γ-/δ-lactones,\nwhich are widely used in fragrances and flavors. Typically, a carbonyl\nreductase mutant <i>Sm</i>CR<sub>M5</sub> from <i>Serratia\nmarcescens</i>, was coimmobilized by covalent binding with <i>Bm</i>GDH, a glucose dehydrogenase capable of recovering and\nrecycling the cofactor NADPH. After immobilization, <i>Sm</i>CR<sub>M5</sub> and <i>Bm</i>GDH exhibited a 8.9-/8.7-fold\nincrease in catalytic efficiency (<i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub>) and a 57-/15-fold increase in half-life\nat 30 °C, respectively. We demonstrated that coimmobilized dual\nenzymes used in a continuous flow reactor showed a higher reaction\nrate and a higher space-time yield (1586 g·L<sup>–1</sup> d<sup>–1</sup>) than free enzymes and immobilized enzymes\nin a batch reaction for the production of (<i>R</i>)-δ-decalactone.\nThis continuous flow reactor can run continuously for more than 650\nh with 99% <i>ee</i> and 80% conversion, and the total volume\nexceeds 1500 reactor volumes. The robustness of this continuous-flow\nimmobilized enzyme system provides a green and efficient method for\nthe synthesis of high value-added chiral chemicals.