Biocatalytic synthesis of 2‐O‐α‐D‐glucopyranosyl‐L‐ascorbic acid using an extracellular expressed α‐glucosidase from Oryza sativa

Background 2‐O‐α‐D‐Glucopyranosyl‐L‐ascorbic acid (AA‐2G) is an important derivative of L‐ascorbic acid (L‐AA), which has the distinct advantages of non‐reducibility, antioxidation, and reproducible decomposition into L‐AA and glucose. Enzymatic synthesis is a preferred method for AA‐2G production over alternative chemical synthesis owing to the regioselective glycosylation reaction. α‐Glucosidase, an enzyme classed into O‐glycoside hydrolases, might be used in glycosylation reactions to synthesize AA‐2G. Main Methods and Major Results Here, an α‐glucosidase from Oryza sativa was heterologously produced in Pichia pastoris GS115 and used for biosynthesis of AA‐2G with few intermediates and byproducts. The extracellular recombinant α‐glucosidase (rAGL) reached 9.11 U mL–1 after fed‐batch cultivation for 102 h in a 5 L fermenter. The specific activity of purified rAGL is 49.83 U mg–1 at 37°C and pH 4.0. The optimal temperature of rAGL was 65°C, and it was stable below 55°C. rAGL was active over the range of pH 3.0–7.0, with the maximal activity at pH 4.0. Under the condition of 37°C, pH 4.0, equimolar maltose and ascorbic acid sodium salt, 8.7 ± 0.4 g L–1 of AA‐2G was synthesized by rAGL. Conclusions and Implications The production of rAGL in P. pastoris was proved to be beneficial in providing enough enzyme and promoting biocatalytic synthesis of AA‐2G. These studies lay the basis for the industrial application of α‐glucosidase. Graphical and Lay Summary 2‐O‐α‐D‐Glucopyranosyl‐L‐ascorbic acid (AA‐2G) is an important industrial derivative of L‐ascorbic acid (L‐AA), which has the distinct advantages of non‐reducibility, antioxidation, and reproducible decomposition into L‐AA and glucose. In this study, the authors characterized an α‐glucosidase from Oryza sativa, which was recombinantly produced in Pichia pastoris GS115, and its potential for AA‐2G production via transglycosylation of L‐AA was investigated. These studies lay the basis for the industrial application of recombinant α‐glucosidase.