Structural analysis of the α-glucosidase HaG provides new insights into substrate specificity and catalytic mechanism
Structural analysis of the α-glucosidase HaG provides new insights into substrate specificity and catalytic mechanism
复制标题
α-葡萄糖苷酶 HaG 的结构分析为底物特异性和催化机制提供了新的见解
DOI：
10.1107/s139900471500721x
复制
发表时间：
2015-06-01
影响因子：
2.2
通讯作者：
Yao, Min
中科院分区：
文献类型：
--
作者：
Shen, Xing;Saburi, Wataru;Yao, Min
alpha-Glucosidases, which catalyze the hydrolysis of the alpha-glucosidic linkage at the nonreducing end of the substrate, are important for the metabolism of alpha-glucosides. Halomonas sp. H11 alpha-glucosidase (HaG), belonging to glycoside hydrolase family 13 (GH13), only has high hydrolytic activity towards the alpha-(1 -> 4)-linked disaccharide maltose among naturally occurring substrates. Although several three- dimensional structures of GH13 members have been solved, the disaccharide specificity and alpha-(1 -> 4) recognition mechanism of alpha-glucosidase are unclear owing to a lack of corresponding substrate- bound structures. In this study, four crystal structures of HaG were solved: the apo form, the glucosyl- enzyme intermediate complex, the E271Q mutant in complex with its natural substrate maltose and a complex of the D202N mutant with d- glucose and glycerol. These structures explicitly provide insights into the substrate specificity and catalytic mechanism of HaG. A peculiar long beta ->alpha loop 4 which exists in alpha-glucosidase is responsible for the strict recognition of disaccharides owing to steric hindrance. Two residues, Thr203 and Phe297, assisted with Gly228, were found to determine the glycosidic linkage specificity of the substrate at subsite + 1. Furthermore, an explanation of the alpha-glucosidase reaction mechanism is proposed based on the glucosyl- enzyme intermediate structure.
