冠状病毒HCoV-229E S1蛋白片段的免疫原性分析

目的表达和纯化HCoV-229E的S1蛋白片段（S1 417-547）,分析其诱导的免疫应答。方法将纯化蛋白免疫小鼠,ELISA检测小鼠血清特异性抗体以及血清IL-4和IFN-γ含量,流式细胞术检测小鼠脾脏T淋巴细胞亚群分布,观察其诱导的免疫应答。结果表达蛋白经金属螯合获得纯化,并诱导小鼠产生了高滴度的抗体。脾脏CD4^＋和CD8^＋比例均升高,CD4^＋/CD8^＋比值下降;免疫鼠血清IFN-γ和IL-4水平显著升高。结论成功构建了HCoV-229E S1蛋白的表达载体,并在BL21（DE3）中得到了高效表达,表达蛋白免疫小鼠后诱导了明显的细胞和体液免疫应答。

Expression of HCoV-229E coronavirus S1 protein and its induction of immune responses

Objective To express and purify the S1 protein of HCoV-229E coronavirus and analyze its induction of immune response. Method S1 gene fragment was synthesized and cloned to pET21α vector.After sequencing,the S1 gene fragment was digested with Nde I and Xho I and inserted into prokaryotic expression vector pET-21α.The recombinant pET21α/ HCoV-229E S1 was transferred into an expression strain E.coli BL21(DE3) and induced by IPTG express fusion protein.The protein was identified by Western blot and the immune response induced in mice. Result The fusion protein of 14.4 kD was expressed mainly as inclusion body. The fragement of S1 protein was confirmed by Western blot.The purified protein induced a moderate degree of immune responses in mice. The percentage of CD8+ T lymphocyte of the splten increased dramatically while the CD4+/CD8+ ratio decreased significantly when compared with mice control.The significant difference of IFN-γ and IL-4 concentration in sera were watched between the immunize and control mice.Conclusion The recombinant S1 protein could be expressed in E.coli BL21 with induction of both humoral and cell mediated immune responses in mice.