重组蛋白AgrC的表达纯化及人工超分子体系的构建

Expression and Purification of Recombinant AgrC and Construction of Its Artificial Supramolecular System

目的:克隆表达金黄色葡萄球菌agr系统的受体蛋白AgrC,构建人工超分子体系。方法:构建基因工程菌E.coli TOP10-pBAD-AgrC,对诱导条件进行优化,对蛋白的提取方法进行分析优化。表达产物使用Western blot鉴定,His-tag镍柱亲和层析纯化。使用肽脂质N+C5Gly2C16为膜材料制备囊泡作为载体,将纯化后的蛋白在囊泡上镶嵌。结果:构建的基因工程菌在18℃,0.002%的阿拉伯糖浓度下具有最高的表达效率,以内膜形式存在的AgrC蛋白在表面活性剂BDH中溶解效率最佳。结论:成功诱导表达并纯化了重组蛋白AgrC,成功利用其构建了人工超分子体系。

Objective： To express and purify AgrC,a receptor protein of Staphylococcus aureus and to construct its artificial supramolecular system.Methods： AgrC expression bacterial strain,E.coli TOP10-pBAD-AgrC was constructed.The induction conditions and the extracting methods of the protein were optimized.The recombinant protein was identified by Western blot and purified with HisTrap affinity column.The vesicle was prepared with peptide lipid N＋C5Gly2C16,and the purified protein was subsequently inset into the vesicle.Results： The constructed engineering bacterium had an optimal expression at 18℃ and with 0.002% arabinose.The protein existing in membrane was maximally dissoluted with BDH.Conclusion： The recombinant AgrC was expressed and purified successfully,and an artificial supramolecular system was constructed.