髓鞘调控因子的镍金属螯合纯化及影响因素研究  

Purification and influencing factors of myelin regulatory factors by nickel metal chelation

作　　者：鄢树枫 孙传龙 陈作宜 YAN Shufeng;SUN Chuanlong;CHEN Zuoyi(Fujian Provincial Medical Plant Erploitation and Utilization Engineering Research Center,Sanming University,Sanming 365004,China)

机构地区：[1]三明学院药用植物开发利用福建省高校工程技术研究中心,福建三明365004

出　　处：《扬州大学学报（农业与生命科学版）》2024年第1期67-74,共8页Journal of Yangzhou University：Agricultural and Life Science Edition

基　　金：福建省自然科学基金资助项目(2020J05092)。

摘　　要：为研究髓鞘调控因子(myelin-gene regulatory factor,MYRF)的镍金属螯合层析纯化方法及其影响因素,以导入MYRF基因的BL21大肠埃希菌为宿主菌,设置不同浓度梯度的咪唑和EDTA洗脱进行镍金属螯合层析柱纯化,通过蛋白质浓度测定、考马斯亮蓝染色和SDS-PAGE分析等方法分析差异,进一步研究蛋白质上清液与镍填料结合时间对MYRF蛋白纯化的影响。结果表明:咪唑洗脱获得的蛋白质浓度与洗脱液浓度呈正比,0.2 mol·L^(-1)的咪唑洗脱液可获得纯度较高的MYRF蛋白;而低浓度(0.05 mol·L^(-1))EDTA洗脱液虽可快速获得高浓度MYRF蛋白,但杂蛋白含量也较高;同时,MYRF蛋白浓度与结合时间呈正相关。综上,MYRF蛋白镍金属螯合层析受洗脱液种类、填料结合时间等多种因素影响,研究结果为MYRF蛋白纯化策略的选择和优化提供参考依据。To study the nickel metal chelation chromatography purification method and influencing factors of myelin gene regulatory factor(MYRF), BL21 E.coli with MYRF gene was used as the host bacterium. Different concentration gradients of imidazole and EDTA were set for nickel metal chelation chromatography column purification. The study was analyzed by protein concentration measurement, coomassie brilliant blue staining, and SDS-PAGE analysis. The effect of binding time between protein supernatant and nickel filler on MYRF protein purification were further studied. The results showed that MYRF concentration obtained by imidazole was proportional to the elution solution concentration, and a 0.2 mol·L^(-1) imidazole solution could obtain MYRF protein with high purity. Although low concentration(0.05 mol·L^(-1)) EDTA can quickly obtain high concentration MYRF protein, the content of impurities was also high. Meanwhile, there was a positive correlation between MYRF concentration and binding time. In summary, MYRF protein nickel metal chelation chromatography is influenced by multiple factors such as the type of eluent and the binding time of fillers, providing reference basis for the selection and optimization of MYRF protein purification strategies.

关 键 词：镍柱 咪唑 EDTA 蛋白质纯化 结合时间 

分 类 号：Q51[生物学—生物化学]