GST与戊型肝炎病毒ORF2编码蛋白融合表达形成的新抗原表位的鉴定  

Identification of a novel antigenic epitope on GST fusion-expressed and ORF2-encoded proteins of hepatitis E virus

作　　者：梁久红[1] 梁立敏[1] 董敏[1] 韩振格[1] 董晨[1] 孟继鸿[1] 

机构地区：[1]东南大学医学院病原生物学与免疫学系,江苏南京210009

出　　处：《细胞与分子免疫学杂志》2008年第4期321-323,327,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家高技术研究发展计划(863)资助项目(2006AA02A235);国家自然科学基金资助项目(30671858);东南大学优秀博士生学位论文资助项目(YBJJ0414)

摘　　要：目的:以戊型肝炎病毒(HEV)ORF2编码的重组蛋白p166为例,研究蛋白标签GST对融合表达的重组蛋白抗原结构的影响。方法:以HEV中国株重组蛋白p166Chn-GST为免疫原,制备单克隆抗体(mAb),与代表HEV4个基因型的摩洛哥株、墨西哥株、美国株和中国株p166的GST或His融合蛋白、中国株非融合重组蛋白p179Chn以及GST融合的HEV无关蛋白进行ELISA检测,鉴定mAb所识别的抗原表位。结果:获得3株稳定分泌抗p166Chn-GST的杂交瘤细胞株,分泌的mAb1A8、9B4和8H10与p166Chn-GST反应,与GST不反应。其中1A8和9B4可与带GST标签的4种p166-GST蛋白以及N和C端截短的p146Chn-GST、p137Chn-GST反应,而不与4种p166-His蛋白反应,也不与p179Chn反应,与HEV病毒颗粒竞争试验阴性,与GST融合的HEV无关蛋白无交叉反应性,表明1A8和9B4识别的抗原表位不是HEV病毒颗粒表面天然存在的抗原表位,而是GST与HEV ORF2编码蛋白的465-601aa区段序列共同形成的新的抗原表位。结论:GST能够赋予基因工程重组蛋白以新的抗原特性,它与融合表达的重组蛋白可以共同形成新的抗原表位。AIM： To investigate the effect of a GST tag on the antigenic structure of GST fusion-expressed and ORF2-encoded recombinant proteins of hepatitis E virus （HEV）. METHODS： The monoclonal antibodies （mAb） were prepared with a GST fusion protein, p166Chn-GST, which was derived from a Chinese HEV strain. Then they were tested by indirect ELISA, competition ELISA and Western blot with different GST fusion, His fusion or non-fusion recombinant proteins derived from HEV reference strains of all 4 genotypes and other non-HEV recombinant proteins. RESULTS： Three mAb named 1A8, 9B4 and 8H10 were obtained. All of them reacted to p166Chn-GST but did not react to GST. mAb 1A8 and 9B4 reacted to 4 p166-GST proteins of different HEV genotypes and 2 N- or C-terminal truncated p166Chn-GST proteins named p146Chn-GST and p137Chn-GST, but they did not react to 4 p166-His proteins of different HEV genotypes and a non-fusion p179Chn protein. No detectable signals were found when 1A8 and 9B4 were subjected to HEV antigen competition ELISA or Western blot after SDS-PAGE. No cress reaction was observed between the two mAb and HEV-irrelevant GST fusion proteins, either. CONCLUSION： A novel antigenic epitope recognized by mAb 1A8 and 9B4 appears on the GST fusionexpressed and ORF2-encoded HEV recombinant proteins and it is dependent on the conformational folding of both GST and HEV sequences.

关 键 词：GST融合表达 戊型肝炎病毒 单克隆抗体 抗原表位 

分 类 号：R373.21[医药卫生—病原生物学]