Whole-cell biosynthesis of 2-O-α-D-glu-copyranosyl-sn-glycerol by recombinant Bacillus subtilis

Abstract:

2-O-α-D-glu-copyranosyl-sn-glycerol is a high value-added product with prospective application in food, cosmetics, health products and pharmaceutical industries. However, industrial scale of 2-O-α-D-glu-copyranosyl-sn-glycerol has not yet been applied in China, and there are few related reports on 2-O-α-D-glu-copyranosyl-sn-glycerol synthesis. The purpose of this experiment is to develop a method for catalyzing the synthesis of food-grade 2-O-α-D-glu-copyranosyl-sn-glycerol using whole cells of “Generally Recognized as Safe” (GRAS) recombinant Bacillus subtilis. In our work, a recombinant B. subtilis 168/pMA5-gtfA that heterologously expressing Leuconostoc mesenteroides sucrose phosphorylase was constructed and used as a whole-cell catalyst to synthesize 2-O-α-D-glu-copyranosyl-sn-glycerol. Optimizing the culture temperature, time and whole cell transformation conditions has increased the yield of 2-O-α-D-glu-copyranosyl-sn-glycerol. The results showed that 1.43 U/mL of sucrose phosphorylase was achieved in B. subtilis 168/pMA5-gtfA after culturing for 20 h at 30 °C in fermentation medium. The highest conversion rate reached 75.1%, and the yield of 2-O-α-D-glu-copyranosyl-sn-glycerol was 189.3 g/L with an average transformation rate of 15.6 mmol/(L·h) after 48 hours whole-cell transformation with the sucrose concentration of 1 mol/L and the glycerol concentration of 2.5 mol/L at 30 °C, OD600 40 and pH 7.0. This is the highest yield of 2-O-α-D-glu-copyranosyl-sn-glycerol synthesized catalytically by recombinant B. subtilis that was ever reported, and this study provides the theoretical and experimental basis for the industrial production and application of 2-O-α-D-glu- copyranosyl-sn-glycerol.