Heterologous Expression of a Thermostable α-Glucosidase from Geobacillus sp. Strain HTA-462 by Escherichia coli and Its Potential Application for Isomaltose⁻Oligosaccharide Synthesis.

Isomaltose-oligosaccharides (IMOs), as food ingredients with prebiotic functionality, can be prepared via enzymatic synthesis using alpha-glucosidase. In the present study, the alpha-glucosidase (GSJ) from Geobacillus sp. strain HTA-462 was cloned and expressed in Escherichia coli BL21 (DE3). Recombinant GSJ was purified and biochemically characterized. The optimum temperature condition of the recombinant enzyme was 65 degrees C, and the half-life was 84 h at 60 degrees C, whereas the enzyme was active over the range of pH 6.0-10.0 with maximal activity at pH 7.0. The alpha-glucosidase activity in shake flasks reached 107.9 U/mL and using 4-Nitrophenyl beta-D-glucopyranoside (pNPG) as substrate, the K-m and Vmax values were 2.321 mM and 306.3 U/mg, respectively. The divalent ions Mn2+ and Ca2+ could improve GSJ activity by 32.1% and 13.8%. Moreover, the hydrolysis ability of recombinant alpha-glucosidase was almost the same as that of the commercial alpha-glucosidase (Bacillus stearothermophilus). In terms of the transglycosylation reaction, with 30% maltose syrup under the condition of 60 degrees C and pH 7.0, IMOs were synthesized with a conversion rate of 37%. These studies lay the basis for the industrial application of recombinant alpha-glucosidase.