人参皂苷Rb3木糖苷酶的纯化及其酶学性质

采用DEAE-Cellulose DE52阴离子交换柱层析的方法对微生物Absidia sp.GRB3-X8r菌产特异性人参皂苷Rb3木糖苷酶进行分离纯化,分离后该酶在SDS-PAGE上呈现单一蛋白质条带,并对其酶学性质进行研究。研究表明:人参皂苷Rb3木糖苷酶的最适反应pH为3.0,在pH2.2~8.0范围内相对稳定;最适温度为40 ℃,在20~60 ℃范围内具有较好的稳定性;金属离子K+、Na+、Mg2+、Mn2+、Ca2+离子对酶反应无明显作用,Zn2+、Pb2+、Ni2+对酶反应有抑制作用。SDS-PAGE电泳结果表明酶蛋白的相对分子量约为66.7 kDa。反应动力学研究结果显示K_m值为65.63 mmol/L,V_max为2.03 mmol/(h·L)。通过对酶的催化特性研究表明,该酶能水解人参皂苷Rb3木糖基,生成人参皂苷Rd。

Purification and Characterization of Ginsenoside Rb3 Xylosidase

A ginsenoside Rb3 xylosidase from ploughshares mildew strains Absidia sp. Grb3-x8r was purified using DEAE-Cellulose DE52 ion exchange column chromatography method. The purified xylosidase showed a single protein band on SDS-PAGE, and its enzymatic properties were studied. The results showed that the optimum reaction conditions pH value of ginsenoside Rb3 xylosyl hydrolase was 3.0, which had relatively stable in the range of pH2.2~8.0. The optimum temperature was 40℃, which had better stability in the range of 20~60℃;Metal ion K+, Na+, Mg2+, Mn2+, Ca2+ ions had no obvious effects on enzyme reaction, and Zn2+, Pb2+, Ni2+ had inhibitory effects on enzyme reaction. The results of SDS-PAGE electrophoresis showed that the molecular weight of enzyme protein was 66.7 kDa. The results of the reaction kinetics showed that the K_m value was 65.63 mmol/L, and V_max was 2.03 mmol/(h·L).The catalytic properties of the enzyme showed that the enzyme could hydrolyze the xyloside of ginsenoside Rb3 into ginsenoside Rd.