广州管圆线虫抗原IF的体外表达系统的建立和分析  

Angiostrongylus Cantonensis:Construction of Expression System in Vitro and Identification of Antigen Gene IF

作　　者：孟锦绣[1] 何蔼[1] 程梅[1] 李素丽[1] 甘明[1] 徐贵峰[1] 李卓雅[1] 詹希美[1] 

机构地区：[1]中山大学中山医学院寄生虫学教研室

出　　处：《中山大学学报（医学科学版）》2006年第6期619-624,共6页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：国家"十五"科技攻关重大项目(2003BA712A03-07);国家自然科学基金资助项目(301708377);广州市科技计划基金资助项目(2005Z3-7561;2006Z3-C7191);广东省医学科研基金资助项目(B2006003);广东省人民医院科学技术研究基金资助项目(Y200532)

摘　　要：【目的】建立广州管圆线虫特异性抗原中间纤维(IF)基因的体外表达系统,研究IF蛋白的结构和功能关系。【方法】将IF基因亚克隆入原核表达载体pET-30a-c(+)并在原核中诱导表达,Westernblot鉴定融合表达产物的免疫特性,用组氨酸(His)亲合层析柱分离纯化表达产物,鉴定表达体系效率。生物信息学分析目的蛋白结构和功能关系。【结果】抗原基因IF在原核中与6个组氨酸进行了融合表达,以可溶性表达为主,并能在该系统中大量、稳定表达,其相对分子质量Mr=48×103,该表达产物具有较好的抗原性,可被大鼠感染血清识别,抗原IF具有典型的中间纤维家族特征,该蛋白具有多个抗原表位存在。【结论】建立了IF的有效原核表达系统,IF抗原蛋白的结构决定了它的属性和功能。[Objective] To construct the expression system in vitro of specific antigen cDNA, the intermediate filament gene （IF） of Angiostrongylus cantonensis, and study IF protein structure correlating its property and function. [Methods] The IF cDNA was subcloned into the prokaryotic expressing vector pET-30a-c（＋） and expressed in vitro. The immuno-characteristics of expressed products were identified by western blot, the expressed products His-tagged were isolated and purified by affinity chromatography resins and the expression ratio was analyzed. The correlation between structure and function of IF proteins was analyzed by bioinformatics. [Results] The antigen IF was expressed as recombinant protein fused with 6 histidine, it could be expressed effectively as soluble protein. The molecular mass of antigen IF was Mr,=48×10^3, it had effective antigenicity and could be recognized by infected rat antiserum. IF protein had the characteristics of intermediate filament protein family and several hydrophilic domain as well. [ Conclusion ] An effective prokaryotic expression system of antigen IF of A ngiostrong＇ylus cantoneruis was constructed, the study showed that the structure of IF protein determine its property and function.

关 键 词：广州管圆线虫 抗原 中间纤维 原核表达 

分 类 号：R383.1[医药卫生—医学寄生虫学]