title: Purification method improvement and characterization of a novel ginsenoside-hydrolyzing beta-glucosidase from Paecilomyces Bainier sp. 229.
abstract: The purification method for a novel ginsenoside-hydrolyzing beta-glucosidase from Paecilomyces Bainier sp. 229 was successfully simplified by the application of microcrystalline cellulose (MCC) as a novel chromatographic matrix. Only two chromatographic steps, Q-Sepharose FF and MCC column in sequence, were required to purify the enzyme to apparent homogeneity. The purified enzyme, with a native molecular weight estimated to be 305 KDa, was composed of three identical subunits of approximately 102 KDa each. The optimal enzyme activity was observed at pH 3.5 at 55 degrees C. It was stable within pH 3-7 and at temperatures lower than 50 degrees C. The optimal substrate for the enzyme was p-nitrophenyl-beta-D-glucoside, followed by ginsenoside Rd, gentiobiose, and ginsenoside Rb1. It converted ginsenoside Rb1 to ginsenoside Rg3 specifically and efficiently. The hydrolyzing pathway of ginsenoside Rb1 by the enzyme was Rb1-->Rd-->Rg3. The specific activities against ginsenoside Rb1 and Rd were 56.7 micromol/min/mg and 129.4 micromol/min/mg respectively.
authors.affiliation: School of Pharmacy, Fudan University, Shanghai 200032, China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Yan
authors.forename: Wei
authors.initials: W
authors.lastname: Zhou
authors.forename: Xingwei
authors.initials: X
authors.lastname: Li
authors.forename: Meiqing
authors.initials: M
authors.lastname: Feng
authors.forename: Pei
authors.initials: P
authors.lastname: Zhou
mesh_terms: D002853:Chromatography, Liquid; D002855:Chromatography, Thin Layer; D004591:Electrophoresis, Polyacrylamide Gel; D036145:Ginsenosides / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008970:Molecular Weight; D010143:Paecilomyces / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D001617:beta-Glucosidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D036145:Ginsenosides; D001617:beta-Glucosidase