高活性大豆异黄酮糖苷酶的分离原料筛选、纯化及性质研究

Selection of Plant Source Used for Separating Isoflavone-glucosidase with High Activity and Its Purification and Properties

通过对32种植物β-葡萄糖苷酶水解染料木苷的活性比较,发现鹰嘴豆β-葡萄糖苷酶具有4.19U/mg的大豆异黄酮糖苷水解酶活性。该酶经硫酸铵分级沉淀、DEAE-Cellulose-52离子交换、Sephadex G-100凝胶层析纯化,纯化了10.1倍,收率为7.2%;SDS-PAGE和Sephadex G-100凝胶层析结果表明,该酶的分子量为73.2kD,含两个亚基;该酶的最适反应温度为45℃;最适pH6.0;当以染料木苷和大豆苷为底物时该酶的米氏常数分别为11.0和19.0μg/ml。温度在30～45℃、pH值在4.5～6.5范围内该酶较稳定:Ag^+、Hg^(2+)和D-葡萄糖酸内酯对该酶有强烈抑制作用。该酶能水解染料木苷和大豆苷,但不能水解纤维二糖和α-乳糖。

The comparison among the activities of isoflavone-glucosidases isolated from 32 species of plants showed that chickpea (Cicer arietinum)β-glucosidase activity is the highest.Then the chickpeaβ-glucosidase was purified by 10.1 folds with the yield of 7.2% after ammonium sulphate precipitation,DEAE-Cellocuse-52 ion exchange and Sephadex G-100 gel filtration chromatography.This purified enzyme has a molecular weight of about 73.2 kD and two subunits.The optimum temperature, pH and K_m towards genistin and daid...