Structural analysis of the α-glucosidase HaG provides new insights into substrate specificity and catalytic mechanism

Xing Shen, Wataru Saburi, Zuoqi Gai, Koji Kato, Teruyo Ojima-Kato, Jian Yu, Keisuke Komoda, Yusuke Kido, Hirokazu Matsui, Haruhide Mori, Min Yao

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

α-Glucosidases, which catalyze the hydrolysis of the α-glucosidic linkage at the nonreducing end of the substrate, are important for the metabolism of α-glucosides. Halomonas sp. H11 α-glucosidase (HaG), belonging to glycoside hydrolase family 13 (GH13), only has high hydrolytic activity towards the α-(1→4)-linked disaccharide maltose among naturally occurring substrates. Although several three-dimensional structures of GH13 members have been solved, the disaccharide specificity and α-(1→4) recognition mechanism of α-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 β→α loop 4 which exists in α-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 α-glucosidase reaction mechanism is proposed based on the glucosyl-enzyme intermediate structure.

Original languageEnglish
Pages (from-to)1382-1391
Number of pages10
JournalActa Crystallographica Section D: Biological Crystallography
Volume71
DOIs
Publication statusPublished - Jan 1 2015
Externally publishedYes

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Keywords

  • glucosyl-enzyme intermediate
  • glycoside hydrolase family 13
  • Halomonas sp. H11
  • reaction mechanism
  • substrate specificity
  • α-glucosidase

ASJC Scopus subject areas

  • Structural Biology

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