H3N2亚型犬流感病毒头部缺失血凝素蛋白的真核表达及抗原性研究

Eukaryotic expression of the headless hemagglutinin of H3N2 canine influenza virus and its antigenicity

为获得一种针对不同亚型流感病毒的共同保护性抗原,本研究以H3N2亚型犬流感病毒(canine influenza virus,CIV)的主要抗原蛋白血凝素(HA)为靶标,以四甘氨酸接头肽替换HA头部结构域,将免疫反应定向至更保守的HA茎部区,同时在HA茎部添加T4折叠三聚体以保护蛋白天然构象,通过杆状病毒表达系统表达头部缺失HA蛋白,并验证其抗原性。结果表明,利用昆虫细胞表达的头部缺失,HA蛋白单一性较好,分子量为47 ku,与CIV全病毒血清抗体可发生特异性结合反应,且头部去除区HA蛋白血清抗体可抑制H1N1和H3N2亚型流感病毒感染所致的细胞病变效应,中和效价分别为1∶160和1∶320。研究结果为制备针对不同亚型流感病毒(如H1N1和H3N2)的广谱疫苗提供了可能。

In order to obtain a common protective antigen against different subtypes of influenza A viruses,hemagglutinin(HA),the main antigen protein of canine influenza virus(CIV)of the H3 N2 subtype,was used as the target in this study.To direct the immune response to the HA stem region,which is more conservative,the head domain was replaced with a tetraglycine linker peptide.Also,a T4 folded trimer motif was inserted into the HA stem to protect the natural conformation of the protein.The HA protein without the head region was expressed through the baculovirus expression system,and its antigenicity was verified.The results showed that the headless HA protein was successfully expressed by insect cells,with a molecular weight of 47 ku.The expressed protein was able to specifically react with the anti-sera against H3 N2 CIV,and the antibody against the headless HA could inhibit the cytopathic effect caused by H1 N1 or H3 N2 virus infection in vitro,with the neutralizing titers of 1∶160 and 1∶320,respectively.This study laid the foundation for the preparation of a universal influenza vaccine to provide broad-spectrum protection against different subtypes of influenza A viruses.