鲎血细胞中脂多糖结合蛋白TALF在大肠杆菌中的表达研究  

Research on TALF Expression in Escherichia coli

作　　者：王东宁[1] 孙向军[2] 张惟杰[1] 吴祥甫[3] 

机构地区：[1]上海交通大学生命科学与技术学院,上海200030 [2]上海交通大学农业与生物技术学院,上海200030 [3]中科院上海生命科学研究院生物化学与细胞学研究所,上海200031

出　　处：《生物工程学报》2004年第4期540-543,共4页Chinese Journal of Biotechnology

摘　　要：TALF(Tachyleusanti lipoposaccharidefactor)对细菌内毒素 (LPS)的核心部分有抑制作用。研究TALFcDNA基因在大肠杆菌中的表达 ,首先将TALFcDNA基因分别插入大肠杆菌表达载体pGEX 4T 2、pET2 2b、pET2 8a中 ,构建重组表达质粒 ,转化于大肠杆菌BL2 1 (DE3)。结果表明克隆于pET2 2b、pET2 8a中的TALFcDNA基因没有表达 ,而融合了GST的TALF基因 (GST TALF)能够在大肠杆菌中表达 ,并形成包涵体。从 1L培养基中可获得 4mg纯度为 91 %的GST TALF融合蛋白。经复性和纯化后的融合蛋白GST TALF几乎检测不到抑菌活性及LPS中和活性 。The expression of cDNA encoding Tachyleus anti lipoposaccharide (LPS) factor, which is of interest for use as a potential inhibitor of the common core subunit of Gram negative bacterial endotoxin. First, the TALF gene was inserted into expression vectors pGEX 4T 2、pET22b and pET28a to construct recombinant expression plasmids. The recombinant plasmids were transformed to E.coli BL21 (DE3) and the expression of TALF was examined. Results show that TALF in pET22b and pET28a vectors can't be expressed. Only the fusion protein GST TALF was expressed in E.coli BL21 existing as inclusion bodies. From 1 liter of culture, about 4mg of fusion protein GST TALF with 91% purity was finally obtained. No apparent bactericidal activity and LPS neutralizing activity of the fusion protein GST TALF were found. After digested with thrombin, the fusion protein GST TALF exhibited strong bactericidal activity and LPS neutralizing activity.

关 键 词：融合蛋白 包涵体 变复性 抑菌活性 LPS中和活性 

分 类 号：R378[医药卫生—病原生物学]