新型佐剂——鞭毛蛋白及其突变体的原核表达和纯化  

Prokaryotic Expression and Purification of Flagellins and Their Mutants as the Neotype Adjuvants

作　　者：吴晓芳[1,2,3] 李树龙[3] 戴红梅[3] 付桂明[1,2] 方宏清[3] 

机构地区：[1]南昌大学食品科学与技术国家重点实验室,江西南昌330047 [2]南昌大学生命科学与食品工程学院,江西南昌330047 [3]军事医学科学院生物工程研究所,北京100850

出　　处：《生物技术通讯》2011年第5期691-695,共5页Letters in Biotechnology

摘　　要：目的:改构鞭毛蛋白可变区,并优化目的蛋白表达与纯化参数,为评价并获得新型高效蛋白佐剂奠定基础。方法:以鼠伤寒沙门菌基因组为模板钓取2种鞭毛蛋白FljB和FliC的编码基因,分别构建FljB和FliC及其对应的删除可变区的突变体BNLC和CNLC的表达载体,在大肠杆菌中诱导表达,并纯化目的蛋白。结果与结论:原初鞭毛蛋白FljB和FliC均以胞内可溶形式表达,而改造后的鞭毛蛋白BNLC和CNLC则以包涵体形式表达。通过优化纯化条件,分别建立了针对4种蛋白的纯化方法,得到无热源、纯度大于95%的目的蛋白,为进一步研究鞭毛蛋白的结构与佐剂效应的关系奠定了基础。Objective： This study for deleting of flagellin＇s central hypervariable domain,optimizating parameters of their protein＇s expression and purification was carried out,and it should contribute to gaining new and high effi-cient adjuvants actively.Methods： Have fished the genes of two flagellins FljB and FliC from genome of Salmonel-la typhimurium,we constructed the expression vectors for FljB and FliC and their relative mutants BNLC and CNLC,which variable regions were deleted selectively,then we induced the proteins expressin in E.coli,and puri-fied these proteins respectively.Results Conclusion： FljB and FliC expressed in dissoluble form in cells,and BNLC and CNLC expressed with inclusion bodies.Through optimizing the purification conditions,we setted up pu-rification methods of four proteins respectively,and getting apyrogenic proteins which purity exceed 95%.It estab-lished a foundation for studying the relationship of structure and adjuvant effiency of flagellin for the next step.

关 键 词：鞭毛蛋白 突变体 原核表达 纯化 

分 类 号：Q78[生物学—分子生物学]