作　　者： ; ; ; ; ;

机构地区： 广东药学院生命科学与生物制药学院生物制药研究所

出　　处： 《广东药学院学报》 2008年第3期282-286,共5页

摘　　要： 目的对严重急性呼吸系统综合征冠状病毒（SARS—CoV）刺突蛋白（Spike，S）S1段进行原核重组表达，并检测表达产物的抗原性。方法对S1中的HLA抗原表位进行预测分析，根据分析结果选定用于重组表达的区段，利用pET21b载体进行原核表达，将产物纯化后利用SARS抗血清进行抗原性检测。结果抗原表位分析表明，S1中包含有多个能够被HLA分子有效提呈的抗原表位，综合考虑表位肽的分布、重组表达的效率、重组蛋白的空间构象以及引物设计的优化等因素，选取S1中的关键区域259～565aa进行原核重组表达，得到了包涵体形式的重组蛋白，经过纯化和复性，获得了纯度较高的可溶性S1蛋白；用SARS抗血清对此重组蛋白进行免疫学鉴定，表明该蛋白具有SARS特异的抗原性。结论重组表达的S1蛋白具有较强的抗原性，为进一步的疫苗研究和抗体筛选奠定了基础。 Objective To express and confirm the antigenicity of the S1 fragment of spike protein of SARS-CoV. Methods The epitopes recognized by HLA in S1 were analyzed, and the proper fragment was selected to express with pET21b vector. The recombinant protein was subject to antigenicity assay after purification. Results Epitope analysis identified the nonamers, which could be presented by HLA molecule, in S1 protein. In view of the distribution of epitopes in S1 protein, the efficiency of recombinant expression, the conformation of recombinant protein, the optimization of primers design, and so on, a fragment （259 -565 aa） was selected to express in prokaryotic cell. The recombinant protein expressed as inclusion body was subject to purifying, denaturation, and renaturation, and purified soluble S1 protein was obtained. The special SARS-CoV antigenicity of this protein was identified by enzyme-linked immunosorbent assay （ELISA） with serum from convalescent SARS patients. Conclusion Recombinant S1 protein showed solid antigenicity. These results laid a foundation for the research of vaccine and neutralizing antibodies.

关 键 词： 蛋白 抗原表位 重组表达

领　　域： [生物学]