Efficient biosynthesis of (S)-1-phenyl-1,2-ethanediol catalyzed by (S)-carbonyl reductase Ⅱ and glucose dehydrogenase

Abstract:

[Objective] To realize efficient biosynthesis of 2-hydroxyacetophenone to (S)-1-phenyl-1,2-ethanediol, we designed a co-expression system containing Candida parapsilosis CCTCC M203011 (S)-carbonyl reductase Ⅱ (SCRⅡ) and Bacillus sp. YX-1 glucose dehydrogenase (GDH) in Escherichia coli BL21(DE3), based on the optimal ratio between the specific activities of the two enzymes. [Methods] The enzymes SCRⅡ and GDH were purified from their corresponding recombinant E. coli strains. When the purified SCRⅡ and GDH were used for the reduction of 2-hydroxyacetophenone to (S)-1-phenyl-1,2-ethanediol, the optimal ratio between their specific activities, the optimal temperature and pH were determined. Based on above results, a co-expression system E. coli BL21(DE3)/S-SD-AS-G harboring SCRⅡ and GDH was constructed. [Results] SCRⅡ and GDH exhibited specific activities of 1.3 U/mg and 13.5 U/mg. When the total enzyme activity was 1 U, the optimal ratio of their activities is between 1:1 and 5:1, and the optimal temperature and pH are 30℃ and 7.0, respectively. So we designed a co-expression system E. coli BL21/S-SD-AS-G, in which the ratio of the SCRⅡ and GDH genes is 1:1. The specific activities of SCRⅡ and GDH are 0.76 U/mg and 0.73 U/mg in the cell-free extracts of E. coli BL21(DE3)/S-SD-AS-G, respectively. The ratio between SCRⅡ and GDH activity is 1:1. Under the optimal conditions, the system showed excellent performance to produce (S)-1-phenyl-1,2-ethanediol with an optical purity and a yield both over 99% during the reduction of 2-hydroxyacetophenone. With respect to the recombinant E. coli BL21(DE3)/pET-SCRⅡ, the co-expression system obviously improved the yield of (S)-1-phenyl-1,2-ethanediol and reduced biotransformation time from 24 h to 13 h. [Conclusion] This work provides the research foundation on the construction of a co-expression system containing a target chiral catalyst and a cofactor-regeneration enzyme for efficient chiral biosynthesis based on the optimal ratio of SCRⅡ and GDH activities.