黑曲霉A9葡萄糖氧化酶的提取纯化

Extraction and Purification of Glucose Oxidase from Aspergillus Niger A9

为研究葡萄糖氧化酶的酶学性质、催化机理等问题,利用黑曲霉(Aspergillus niger)A9,经发酵、破碎细胞,得到胞内葡萄糖氧化酶,然后经超滤浓缩、硫酸铵盐析、DEAE-纤维素离子交换层析、Sephadex G-100凝胶过滤分离纯化。试验结果表明:通过控制离子交换层析与分子筛层析的具体操作条件,大部分杂蛋白被除去,所得酶蛋白经PAGE电泳鉴定为单一条带,达到电泳纯。测定其比活力为1779.29U/mg,比初始发酵液提高20.13倍。建立了一套操作简单、快速且费用低廉的黑曲霉A9葡萄糖氧化酶的提纯方案。

In order to research the properties and catalytic mechanism of glucose oxidase, intracellular glucose oxidase was obtained by fermenting Aspergillus niger A9 and breaking up the mycelium. Glucose oxidase was purified by using ultrafihration, ammonium sulfate fractionation, DEAE-cellulose ion-exchange chromatography and Sephadex G-100 gel filtration. The result indicated that most miscellaneous protein had been removed by controlling the detail conditions of ion-exchange chromatography and molecular sieve chromatography. The glucose oxidase showed a single band in PAGE electrophoresis. The specific activity reached 1 779.29 U/mg and was 20.13 folds than the crude fermenting liquor. This research established a set of method which is easy, fast and inexpensive for extracting and purifing the glucose oxidase from Aspergillus niger A9.