2种阴离子交换柱对α-葡萄糖苷酶的分离纯化效果

Effects of two kinds of anion exchange chromatography in the separation and purification of α-glucosidase

【目的】比较Q-Sepharose Fast Flow和DE-52 2种阴离子交换柱对1株脂环酸芽孢杆菌Alicycloba-cillus contaminans发酵产生的α-葡萄糖苷酶的分离纯化效果,为α-葡萄糖苷酶的规模化生产提供技术支持。【方法】采用硫酸铵分级沉淀法对发酵液中的蛋白质进行沉淀,以分离纯化后的蛋白质含量和酶活为指标,对2种阴离子交换柱的分离纯化效果进行评价。【结果】经过Q-Sepharose Fast Flow阴离子交换层析柱分离得到的α-葡萄糖苷酶有1个酶活吸收峰,最大酶活为6.99×103 U,在这个吸收峰内蛋白质的含量稳定在26.25μg/mL左右;经过DE-52阴离子交换层析柱分离,α-葡萄糖苷酶的酶活范围较大,最大酶活为5.56×103 U,在酶活吸收峰内,蛋白质含量为21.25～232.5μg/mL,变化范围较大。【结论】Q-Sepharose Fast Flow阴离子交换层析柱对α-葡萄糖苷酶的分离效果优于DE-52层析柱。

【Objective】 The research focused on the comparison between Q-Sepharose Fast Flow and DE-52 in the separation and purification of α-glucosidase from Alicyclobacillus contaminans which was separated from orchard of Shaanxi Province,in order to provide technical support for industrial-scale production of α-glucosidase.【Method】 Firstly,ammonium sulfate was used to deposit protein by different saturation levels,then the concentrated solution was treated by the two kinds of anion exchange chromatography.After that,protein content and enzyme activity were used to evaluate the effects of the two kinds of anion exchange chromatography in the separation of α-glucosidase.【Result】 The results showed one absorption peak of α-glucosidase appeared and the enzyme activity was 6.99×103 U after the treatment of Q-Sepharose Fast Flow,and the protein content was steady at about 26.25 μg/mL in this absorption peak;the range of absorption peak was large after treated by DE-52 and the highest point enzyme activity was 5.56×103 U,but the protein content fluctuated from 21.25 to 232.5 μg/mL in this absorption peak,varying in a large range.【Conclusion】 Anion exchange chromatography Q-Sepharose Fast Flow is better than DE-52 in the separation of α-glucosidase.