南方农业学报2012，Vol.43Issue(8)：1090-1093,4.DOI:10.3969/j:issn.2095-1191.2012.08.1090
类芽孢杆菌胞外胆固醇氧化酶的分离纯化
Isolation and purification of extracellular cholesterol oxidase from Paenibacillus
摘要
Abstract
[Objective]The present study was conducted to investigate the isolation and purification method of extracellular cholesterol oxidase (COD) from Paenibacillus to provide technical support for Paenibacillus ' production and application. [Method]The fermentation of Paenibacillus was treated through circumrotate evaporation evaporation,sucrose diallysis,DEAE-cellulose ion exchange column and Sephadex-G75 gel filtration column chromatography to purify target protein and determining its fundamental enzymological characteristics. [ Result ] After Paenibacillus fermentation broth was processed by DEAE-cellulose ion exchange column and Sephadex-G75 gel filtration column chromatography,the purified enzyme specific activity reached 6.83 U/mg,the recycling rate reached 35.6% and purification multiple was 13.4 times. The purified enzyme demonstrated to be a single band on SDS-PAGE,and its molecular weight was about 58 kDa. Applied to cholesterol in different concentrations,Km of the purified COD was 3.3×l0-5 mol/L using Lineweaver Burk mapping. The result of thin layer chromatography indicated that the product was cholesteric-4-olefin-3-ketone and the purified enzyme was COD. [ Conclusion ]Electrophoretical purity extracellular COD from Paenibacillus was obtained through isolation and purification,and it is worthy of more application and studies for its positive affinity to cholesterol.关键词
类芽孢杆菌/胆固醇氧化酶/分离/纯化Key words
Paenibacillus/cholesterol oxidase/isolation/purification分类
化学化工引用本文复制引用
杨辉,吕彦茹,余磊,赵芳,王发合,李杰民,梁海秋,朱萍,周河治..类芽孢杆菌胞外胆固醇氧化酶的分离纯化[J].南方农业学报,2012,43(8):1090-1093,4.基金项目
广西自然科学基金项目(桂科自0728017) （桂科自0728017）
