α-葡萄糖苷合成酶的制备及其合成低聚糖的研究  

The Study on the Construction of α-Glycosynthase and the Synthesis of Oligosaccharides

作　　者：毕云枫[1] 姜仁凤[1] 王丽娜[2] 徐琳琳[1] 姜珊[1] 王玉华[1] 

机构地区：[1]吉林农业大学食品科学与工程学院,长春130118 [2]吉林医药学院药学院,吉林132013

出　　处：《食品工业》2017年第5期162-166,共5页The Food Industry

基　　金：吉林省教育厅"十二五"科学技术研究项目(吉教科合字[2015]第207号);吉林省科技厅科技创新人才培育计划项目(项目编号:20140519011JH);吉林省科技厅科技成果转化计划项目(项目编号:20130302017NY)

摘　　要：一种新型的合成低聚糖的方法是利用来自于嗜热古菌Thermotoga neapolitana的α-葡萄糖苷酶及其突变后形成的糖苷合成酶。这种糖苷合成酶的构建是在糖苷酶催化中心亲核体氨基酸位点用非亲核体氨基酸取代亲核体氨基酸,从而使糖苷酶水解活性丧失,只具有糖苷合成能力,使寡糖的合成量增加。此糖苷合成酶合成寡糖过程是利用氟代吡喃葡萄糖作为供体,麦芽糖作为受体,产物为麦芽三糖。产物用HPLC进行分析,然后通过正交试验对合成低聚糖反应条件进行优化,得到最适加酶量为0.06 mg/mL,最适底物浓度为35 mmol/L,最适反应时间为6 h。A novel method for the synthesis of oligosaccharides was used to form the Thermotoga neapolitana a-glucosidase and the glycosynthase by mutagenesis. The glycosynthases construction is in glycosidase catalysed center nucleophilic amino acid sites with non nucleophilic amino acid substitution nucleophilic amino acid, and then makes the hydrolysis activity lost; Only having the ability of glycoside synthesis, so that the oligosaccharides increased. The glycosynthases transglycosylation reaction is used fluoro glucopyranose as donor, with maltose as the acceptor, the product is maltotriose. The products were analyzed by HPLC, and the optimal reaction conditions were optimized by orthogonal experiments, the optimum enzyme amount was 0.06 mg/mL, the optimum substrate concentration was 35 mmol/L, the optimum reaction time was 6 h.

关 键 词：Α-葡萄糖苷酶 低聚糖合成 定点突变 

分 类 号：TS201.2[轻工技术与工程—食品科学]