Efficient production of (S)-1-phenyl-1,2-ethanediol using xylan as co-substrate by a coupled multi-enzyme Escherichia coli system

Abstract

Background: (S)-1-phenyl-1,2-ethanediol is an important chiral intermediate in the synthesis of liquid crystals and chiral biphosphines. (S)-carbonyl reductase II from Candida parapsilosis catalyzes the conversion of 2-hydroxyacetophenone to (S)-1-phenyl-1,2-ethanediol with NADPH as a cofactor. Glucose dehydrogenase with a Ala258Phe mutation is able to catalyze the oxidation of xylose with concomitant reduction of NADP + to NADPH, while endo-β-1,4-xylanase 2 catalyzes the conversion of xylan to xylose. In the present work, the Ala258Phe glucose dehydrogenase mutant and endo-β-1,4-xylanase 2 were introduced into the (S)-carbonyl reductase II-mediated chiral pathway to strengthen cofactor regeneration by using xylan as a naturally abundant co-substrate. Results: We constructed several coupled multi-enzyme systems by introducing (S)-carbonyl reductase II, the A258F glucose dehydrogenase mutant and endo-β-1,4-xylanase 2 into Escherichia coli. Different strains were produced by altering the location of the encoding genes on the plasmid. Only recombinant E. coli/pET-G-S-2 expressed all three enzymes, and this strain produced (S)-1-phenyl-1,2-ethanediol from 2-hydroxyacetophenone as a substrate and xylan as a co-substrate. The optical purity was 100% and the yield was 98.3% (6 g/L 2-HAP) under optimal conditions of 35 °C, pH 6.5 and a 2:1 substrate-co-substrate ratio. The introduction of A258F glucose dehydrogenase and endo-β-1,4-xylanase 2 into the (S)-carbonyl reductase II-mediated chiral pathway caused a 54.6% increase in yield, and simultaneously reduced the reaction time from 48 to 28 h. Conclusions: This study demonstrates efficient chiral synthesis using a pentose as a co-substrate to enhance cofactor regeneration. This provides a new approach for enantiomeric catalysis through the inclusion of naturally abundant materials.

Microbial alcohol dehydrogenases: recent developments and applications in asymmetric synthesis

“…This multienzyme combination under optimal reaction conditions successfully produced (S)-1phenyl-1,2-ethanediol with 100% optical purity in 98.3% yield. 88 A multienzyme system was prepared using the S. cerevisiae spore encapsulation method. The biocatalytic system involved a mutant (S)-specific carbonyl reductase II (SCR II) (Glu228Ser) from C. parapsilosis and Bacillus sp.…”

“…This multienzyme combination under optimal reaction conditions successfully produced ( S )-1-phenyl-1,2-ethanediol with 100% optical purity in 98.3% yield. 88…”

Construction of Bi-Enzyme Self-Assembly Clusters Based on SpyCatcher/SpyTag for the Efficient Biosynthesis of (R)-Ethyl 2-hydroxy-4-phenylbutyrate

“…To determine the optimal pH, the reduction in OPBE by BESCs was assayed in 100 mM PB at different pH levels (5)(6)(7)(8)(9). The prepared BESCs were fully dissolved in buffers with different pH values at a final concentration of 25 µM.…”

“…However, the reaction catalyzed by carbonyl reductase is limited by cofactor recycling. To enhance the regeneration of cofactors in the biosynthesis of chiral alcohols, the transformation efficiency can be improved using multi-enzyme systems, substrate coupling, and enzyme coupling techniques [8][9][10]. In this study, the cofactor NADPH was externally added as a hydrogen donor for the carbonyl reductase-mediated asymmetric reduction of OPBE to promote conversion efficiency and affect the conversion of reduction.…”

Catalytic Synthesis of (S)-CHBE by Directional Coupling and Immobilization of Carbonyl Reductase and Glucose Dehydrogenase

“…However, nicotinamide coenzymes are expensive, unstable, and difficult to reuse, resulting in high costs and limiting their industrial applications [13,14]. In order to optimize the efficiency of coenzymes and improve the biosynthesis of chiral alcohols, researchers introduced advanced techniques such as multi-enzyme systems, substrate coupling, and enzyme coupling [15][16][17]. Therefore, we planned to utilize glucose dehydrogenase (GDH) to regenerate NADPH to provide hydrogen atoms for the carbonyl reductase-mediated COBE asymmetric reduction reaction.…”