食品科学 ›› 2009, Vol. 30 ›› Issue (1): 164-168.doi: 10.7506/spkx1002-6630-200901039

• 生物工程 • 上一篇    下一篇

高活性大豆异黄酮糖苷酶的筛选、纯化及性质研究

潘利华,罗建平* ,罗水忠,徐学玲,王贵娟   

  1. 合肥工业大学生物与食品工程学院
  • 收稿日期:2007-12-04 修回日期:2008-07-28 出版日期:2009-01-01 发布日期:2010-12-29
  • 通讯作者: 潘利华 E-mail:llihuapan@163.com, panlihua612@yahoo.com.cn
  • 基金资助:

    安徽省自然科学基金项目(050410301);安徽省高校青年教师资助计划项目(2006jp1022)

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

PAN Li-hua,LUO Jian-ping*,LUO Shui-zhong,XU Xue-ling,WANG Gui-juan   

  1. (School of Biotechnology and Food Engineering, Hefei University of Technology, Hefei 230009, China)
  • Received:2007-12-04 Revised:2008-07-28 Online:2009-01-01 Published:2010-12-29
  • Contact: PAN Li-hua E-mail:llihuapan@163.com, panlihua612@yahoo.com.cn

摘要:

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1.0 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+、Hg2+ 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。

关键词: 大豆异黄酮糖苷水解酶, β-葡萄糖苷酶, 染料木苷, 鹰嘴豆, 酶学性质

Abstract:

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 chickpea β-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 Km towards genistin and daidzin of this enzyme are 45 ℃, 6.0, 11.0 μg/ml and 19.0 μg/ml, respectively. This β- glucosidase is stable at pH and temperature range of 4.0 to 6.5 and 30 to 45 ℃ respectively. Furthermore, this enzyme can effectively hydrolyse genistin and daidzin, but be strongly inhibited by Ag+, Hg2+ and D-glucono-δ-lactone at a concentration of 1 mmol/L.

Key words: soybean isoflavone-glucosidase, β-glucosidase, genistin, Cicer arietinum, enzymatic properties

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