Purification and characterization of two novel beta-galactosidases from Lactobacillus reuteri.

The intracellular beta-galactosidase (beta-gal) enzymes from two strains of Lactobacillus reuteri, L103 and L461, were purified by ammonium sulfate fractionation, hydrophobic interaction, and affinity chromatography. Both enzymes are heterodimers with a molecular mass of 105 kDa, consisting of a 35 kDa subunit and a 72 kDa subunit. Active staining of L. reuteri L103 and L461 beta-gal with 4-methylumbelliferyl beta-d-galactoside showed that the intact enzymes as well as the larger subunits possess beta-galactosidase activity. The isoelectric points of L. reuteri L461 and L103 beta-gal were found to be in the range of 3.8-4.0 and 4.6-4.8, respectively. Both enzymes are most active in the pH range of 6-8; however, they are not stable at pH 8. The L. reuteri beta-galactosidases are activated by various mono- and divalent cations, including Na(+), K(+), and Mn(2+), and are moderately inhibited by their reaction products d-glucose and d-galactose. Because of their origin from beneficial and potentially probiotic lactobacilli, these enzymes could be of interest for the synthesis of prebiotic galacto-oligosaccharides.