青春双歧杆菌DSM20083中两种α-葡萄糖苷酶的克隆与特性分析
Cloning and characterization of two alpha-glucosidases from Bifidobacterium adolescentis DSM20083.
作者信息
van den Broek L A M, Struijs K, Verdoes J C, Beldman G, Voragen A G J
机构信息
Laboratory of Food Chemistry, Wageningen University, P.O. Box 8129, 6700 EV Wageningen, The Netherlands.
出版信息
Appl Microbiol Biotechnol. 2003 Mar;61(1):55-60. doi: 10.1007/s00253-002-1179-1. Epub 2003 Jan 25.
Two alpha-glucosidase encoding genes (aglA and aglB) from Bifidobacterium adolescentis DSM 20083 were isolated and characterized. Both alpha-glucosidases belong to family 13 of the glycosyl hydrolases. Recombinant AglA (EC 3.2.1.10) and AglB (EC 3.2.1.20), expressed in Escherichia coli, showed high hydrolytic activity towards isomaltose and pnp-alpha-glucoside. The K(m) for pnp-alpha-glucoside was 1.05 and 0.47 mM and the V(max) was 228 and 113 U mg(-1) for AglA and AglB, respectively. Using pnp-alpha-glucoside as substrate, the pH optimum for AglA was 6.6 and the temperature optimum was 37 degrees C. For AglB, values of pH 6.8 and 47 degrees C were found. AglA also showed high hydrolytic activity towards isomaltotriose and, to a lesser extent, towards trehalose. AglB has a high preference for maltose and less activity towards sucrose; minor activity was observed towards melizitose, low molecular weight dextrin, maltitol, and maltotriose. The recombinant alpha-glucosidases were tested for their transglucosylation activity. AglA was able to synthesize oligosaccharides from trehalose and sucrose. AglB formed oligosaccharides from sucrose, maltose, and melizitose.
从青春双歧杆菌DSM 20083中分离并鉴定了两个编码α-葡萄糖苷酶的基因（aglA和aglB）。这两种α-葡萄糖苷酶均属于糖基水解酶家族13。在大肠杆菌中表达的重组AglA（EC 3.2.1.10）和AglB（EC 3.2.1.20）对异麦芽糖和对硝基苯基-α-葡萄糖苷表现出高水解活性。AglA和AglB对硝基苯基-α-葡萄糖苷的K(m)分别为1.05和0.47 mM，V(max)分别为228和113 U mg(-1)。以对硝基苯基-α-葡萄糖苷为底物时，AglA的最适pH为6.6，最适温度为37℃。对于AglB，最适pH值为6.8，最适温度为47℃。AglA对异麦芽三糖也表现出高水解活性，对海藻糖的水解活性较低。AglB对麦芽糖有高度偏好，对蔗糖的活性较低；对蜜二糖、低分子量糊精、麦芽糖醇和麦芽三糖的活性较低。对重组α-葡萄糖苷酶的转糖基化活性进行了测试。AglA能够从海藻糖和蔗糖合成寡糖。AglB能从蔗糖、麦芽糖和蜜二糖形成寡糖。
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