Cloning, Expression, Purification, and Characterization of beta-Galactosidase from Bifidobacterium longum and Bifidobacterium pseudocatenulatum

Expression and purification of beta-galactosidases derived from Bifidobacterium provide a new resource for efficient lactose hydrolysis and lactose intolerance alleviation. Here, we cloned and expressed two beta-galactosidases derived from Bifidobacterium. The optimal pH for BLGLB1 was 5.5, and the optimal temperature was 45 degrees C, at which the enzyme activity of BLGLB1 was higher than that of commercial enzyme E (300 +/- 3.6 U/mg) under its optimal conditions, reaching 2200 +/- 15 U/mg. The optimal pH and temperature for BPGLB1 were 6.0 and 45 degrees C, respectively, and the enzyme activity (0.58 +/- 0.03 U/mg) under optimum conditions was significantly lower than that of BLGLB1. The structures of the two beta-galactosidase were similar, with all known key sites conserved. When o-nitrophenyl-beta-D-galactoside (oNPG) was used as an enzyme reaction substrate, the maximum reaction velocity (V-max) for BLGLB1 and BPGLB1 was 3700 +/- 100 U/mg and 1.1 +/- 0.1 U/mg, respectively. The kinetic constant (K-m) of BLGLB1 and BPGLB1 was 1.9 +/- 0.1 and 1.3 +/- 0.3 mmol/L, respectively. The respective catalytic constant (k(cat)) of BLGLB1 and BPGLB1 was 1700 +/- 40 s(-1) and 0.5 +/- 0.02 s(-1), respectively; the respective k(cat)/K-m value of BLGLB1 and BPGLB1 was 870 L/(mmol.s) and 0.36 L/(mmol.s), respectively. The K-m, k(cat) and V-max values of BLGLB1 were superior to those of earlier reported beta-galactosidase derived from Bifidobacterium. Overall, BLGLB1 has potential application in the food industry.