PURIFICATION AND CHARACTERIZATION OF BETA-D-GLUCOSIDASE (BETA-D-FUCOSIDASE) FROM BIFIDOBACTERIUM BREVE CLB ACCLIMATED TO CELLOBIOSE

The beta-D-glucosidase (EC, 3.2.1.21) activity of Bifidobacterium brev e 203 was increased by acclimation with cellobiose, and the enzyme was purified to homogeneity from cell-free extracts of an acclimatized st rain of B. breve clb, by ammonium sulfate fractionation and column chr omatographies of anion-exchange, gel filtration, Gigapaite, and hydrop hobic interaction, This enzyme had not only beta-D-glucosidase activit y but also beta-D-fucosidase activity, which is specific to Bifidobact eria in intestinal flora, The molecular weight of the purified enzyme was estimated to be 47,000-48,000 and the enzyme was assumed to be a m onomeric protein, The optimum pH and temperature of the enzyme were ar ound 5.5 and 45 degrees C, respectively. The enzyme was stable up to 4 0 degrees C and between pH 5 and 8, The isoelectric point of the enzym e was 4.3 and the K-m values for p-nitrophenyl-beta-D-glucoside and p- nitrophenyl-beta-D-fucoside were 1.3 mM and 0.7 mM, respectively, This enzyme had also transferase activity for the beta-D-fucosyl group but not for the beta-D-glucosyl group, The N-terminal amino acid sequence of this enzyme was similar to those of beta-D-glucosidase from other bacteria, actinomycetes, and plants.