GFE-1多肽与重组人肿瘤坏死因子α融合蛋白的构建及其体外活性和体内分布研究  

Construction, Bioactivity and Distribution of GFE-1-rmhTNF Fu sion Protein

作　　者：周治中[1,2] 李维娜[1,2] 张存[1,2] 刘楠楠[3] 张伟 张英起[1,2] 

机构地区：[1]肿瘤生物学国家重点实验室 [2]第四军医大学药学系生物制药学教研室,陕西西安710032 [3]第四军医大学基础部教学实验中心,陕西西安710032

出　　处：《生物技术通讯》2013年第2期178-182,共5页Letters in Biotechnology

摘　　要：目的:构建GFE-1多肽与重组人肿瘤坏死因子α(rmhTNF-α)融合蛋白(GFE-1-rmhTNF),研究该融合蛋白的体外活性和体内分布。方法:利用基因工程方法,将人工合成的编码GFE-1的寡核苷酸片段连接在rmhTNF-α序列的3'端,转入大肠杆菌中诱导表达,采用Q-Sepharose FF阴离子层析柱和SP-Sepharose FF阳离子层析柱纯化蛋白,SDS-PAGE和Western印迹鉴定,测定该融合蛋白的体外活性,观察其在小鼠体内的分布情况。结果:构建了融合蛋白GFE-1-rmhTNF,并在大肠杆菌中获得高效表达。体外活性实验显示,GFE-1-rmhTNF对L929细胞有明显的杀伤活性;体内分布实验证实,GFE-1-rmhTNF在小鼠肺组织的富集远高于肝肾组织。结论:构建了融合蛋白GFE-1-rmhTNF,可显著杀伤L929细胞并特异性富集于小鼠肺组织。Objective： To construct GFE-I-rmhTNF fusion protein and study its bioactivity in vitro and distribu- tion in vivo. Methods： The synthetic GFE-I oligonucleotide fragment was linked to the 3 terminal of the se- quence encoding rmhTNF-a and expressed in E.coli DH5a. The fusion protein was purified by ion-exchange chro- matography and identified by SDS-PAGE and Western blot. Bioactivity in vitro and distribution in vivo of the fu- sion protein were further analysed. Results： GFE-I-rmhTNF was constructed and expressed in E.coli efficiently. In vitro, its significant cytotoxic activity in L929 cells was observed. In vivo, GFE-I-rmhTNF was dramatically en- riched in the lung tissue of mice than the liver and kidney tissue. Conclusion： We successfully constructed GFE-I-rmhTNF fusion protein. It could kill L929 cells efficiently and specifically enriched in the lung tissue of mice.

关 键 词：人肿瘤坏死因子 GFE-1多肽 融合蛋白 体外活性 体内分布 

分 类 号：R977.4[医药卫生—药品] Q78[医药卫生—药学]