摘要
目的探寻诱导谷胱甘肽转移酶(GST)与Cav1.2钙通道片段CT1重组的易形成包涵体的大分子融合蛋白的提取与纯化的方法。方法在E.coli BL21中转化入p GEX-6p-3/CT1重组质粒并诱导表达,采用GST包涵体变性复性试剂盒和非离子去污剂B-PER分离纯化GST-CT1融合蛋白,用Pull down assay方法鉴定GST-CT1融合蛋白及其生物活性。结果应用GST包涵体变性复性试剂盒和非离子去污剂B-PER能够分离得到高纯度的易形成包涵体的GST-CT1融合蛋白,且分离纯化的GST-CT1融合蛋白具有与钙调蛋白结合的生物活性。结论操作简易的GST包涵体变性复性法能够针对易形成包涵体的GST-CT1融合蛋白进行分离和纯化,且得到的蛋白具有生物学活性。
Objective To explore methods for the extraction and purification of the inclusion body of fusion protein of glutathione transferase （GST） and CT1 fragment of Cav1.2, and to investigate its biologic activity. Methods PGEX-6p-3 recombinant plasmid was transfected into E.coli BL21 to express. GST-CT1 fusion protein with GST inclusion body was purified by solubilization and renaturation kit and B-PER. The biologic activi- ty of GST-CT1 fusion protein was identified by pull down assay. Results The GST-CT1 fusion protein, which is easy to form inclusion body, was successfully purified by the established method and pull down assay showed that the purified protein can bind to calmodulin protein. Conclusion The GST-CT1 fusion protein was successfully purified by denaturation and renaturation method and also showed its normal biologic activity.
出处
《中国医科大学学报》
CAS
CSCD
北大核心
2014年第9期777-780,共4页
Journal of China Medical University
基金
国家自然科学基金(31071004
81100108)
关键词
包涵体
变性再复性
钙离子通道
inclusion body
denaturation and renaturation
calcium channel
