Purification and Characterization of Ginsenoside Ra-Hydrolyzing β-D-Xylosidase from Bifidobacterium breve K-110, a Human Intestinal Anaerobic Bacterium

Abstract

β-D-Xylosidase (EC 3.2.1.37) has been purified from ginsenoside Ra-metabolizing Bifidobacterium breve K-110, which was isolated from human intestinal microflora. β-D-Xylosidase was purified to apparent homogeneity by a combination of ammonium sulfate precipitation, QAE-cellulose, butyl-toyopearl, hydroxyapatit and Q-Sepharose column chromatographies with the final specific activity of 51.8 μmol/min/mg. Molecular weight of β-D-xylosidase is 49 kDa by SDS-PAGE and gel filtration, which consisted of a single subunit. β-D-Xylosidase showed optimal activity at pH 5.0 and 37 °C. The purified enzyme was potently inhibited by PCMS. β-D-Xylosidase acted to the greatest extent on p-nitrophenyl-β-D-xylopyranoside, followed by ginsenoside Ra1 and ginsenoside Ra2. This enzyme hydrolyzed xylan to xylose, but did not act on p-nitrophenyl-β-glucopyranoside, p-nitrophenyl-β-galactopyranoside or p-nitrophenyl-β-D-fucopyranoside. These findings suggest that this is the first reported purification of ginsenoside-hydrolyzing β-D-xylosidase from an anaerobic Bifidobacterium sp.