A new GH13 α-glucosidase from alkaliphilic Bacillus pseudofirmus 703 with both exo-α-l, 4-glucosidase and oligo-l, 6-glucosidase activities toward amylopectin.

Debranching of (1, 6)-α-linkages in starch is of great significance as it is widely used in different industries. In recent years, identifying a single potential enzyme that could debranch (1, 6)-α-linkages and cleave (1, 4)-α-linkages brought great interest as amylopectin possessed both (1, 4)-α- and (1, 6)-α-linkages. In the present study, a 64\,kDa exo-α-l, 4-glucosidase from Bacillus pseudofirmus 703, annotated as Amy112, was cloned and characterized. Biochemical analysis indicated that heterologous Amy112 expressed in Escherichia coli exhibited a high activity against amylopectin, with the optimal temperature and pH of 40°C and pH 7.0, respectively. Addition of K+ ions improved the amy112 activity by 12%, but Li+, Ca2+ and Mg2+ ions showed no significant effect. Amy112 sequence homology revealed that it belonged to glycoside hydrolase family 13, showing 65% identity with α-glucosidase GSJ from Geobacillus sp. HTA-462. This is the first report indicating that Amy112 from B. pseudofirmus 703 belongs to GH13 enzyme family, having both exo-α-1, 4-glucosidase and oligo-l, 6-glucosidase activities. However, no transglycosylation activity was detected in LC-MS analysis. Amy112 would be of great significance of being utilized to debranch starch in different industries in a cost effective manner.