To investigate a novel β-glucosidase from Bifidobacterium breve ATCC 15700 (BbBgl) to produce compound K (CK) via ginsenoside F2 by highly selective and efficient hydrolysis of the C-3 glycoside from ginsenoside Rd, the BbBgl gene was cloned and expressed in E. coli BL21. The recombinant BbBgl was purified by Ni-NTA magnetic beads to obtain an enzyme with specific activity of 37 U/mg protein using pNP-Glc as substrate. The enzyme activity was optimized at pH 5.0, 35°C, 2 or 6 U/ml, and its activity was enhanced by Mn2+ significantly. Under the optimal conditions, the half-life of the BbBgl is 180 h, much longer than the characterized β-glycosidases, and the Km and Vmax values are 2.7 mM and 39.8 µmol/mg/min for ginsenoside Rd. Moreover, the enzyme exhibits strong tolerance against high substrate concentration (up to 40 g/l ginsenoside Rd) with a molar biotransformation rate of 96% within 12 h. The good enzymatic properties and gram-scale conversion capacity of BbBgl provide an attractive method for large-scale production of rare ginsenoside CK using a single enzyme or a combination of enzymes.
Keywords: Bifidobacterium breve; biotransformation; compound K; ginsenoside Rd; β-Glucosidase.