Production of gentiooligosaccharide by recombinant β-glucosidase

Abstract:

Abstract: [Objective] β-glucosidase can be used to prepare gentiooligosaccharide from glucose. The purpose of this study is to obtain β-glucosidase through DNA recombinant technology as well as to optimize the production of gentiooligosaccharide by the recombinant β-glucosidase. [Methods] We cloned bgl, the gene encoding β-glucosidase from Aspergillus niger (CMI CC 324626) into the expression vector pPIC9K to construct the recombinant plasmid pPIC9K- bgl. The vector was then transformed into Pichia pastoris KM71 for extracellular overproduction of β-glucosidase. The activity of the expressed enzyme was measured by the assay of transglucosidation reaction and the transglucosidation product was identified by HPLC and LC-MS. Furthermore, the condition for prepare gentiooligosaccharide by this recombinant β-glucosidase is optimized.[Results] A. niger β-glucosidase was successfully expressed in P. pastoris and the recombinant produced gentiooligosaccharide from glucose. In addition, the main operation parameters of this enzymatic conversion were optimized. At 80% glucose, 60℃, pH 4.5, 1 mmol/L K+, 60 U β-glucosidase per gram substrate, and 48 h reaction time, the gentiooligosaccharide produced reached 50 g/L. [Conclusion] This is the first report of producing gentiooligosaccharide by recombinant β-glucosidase.