扎伊尔埃博拉病毒糖蛋白泛MHC Ⅱ限制性表位预测及验证

Prediction and verification of pan-MHCⅡrestricted epitopes in the glycoprotein of Zaire Ebola virus

目的生物信息学预测获得埃博拉病毒(EBOV)糖蛋白泛主要组织相容性复合体Ⅱ(MHCⅡ)限制性优势表位,分析其特征并筛选鉴定可用于疫苗研制的候选对象。方法通过IEDB分析预测EBOV糖蛋白的优势表位,并对优势表位进行聚类分析,保守性分析和分子对接。同时,利用编码糖蛋白DNA的质粒pVAX-GP免疫小鼠,酶联免疫斑点试验(ELISpot)验证预测获得的表位。结果在构成人群覆盖率达99%的人类白细胞抗原Ⅱ(HLAⅡ)超家族限制性15表位肽预测中筛选获得19个优势表位,H-2筛选获得13个优势表位。ELISpot结果显示pVAX-GP可诱导小鼠脾细胞产生针对合成优势表位的细胞免疫反应,其中表位LFLRATTELRTFSIL和GYYSTTIRYQATGFG在C3H小鼠体内具有显著激活细胞免疫反应的效果。保守性分析结果中存在种间和种内均保守的优势表位,不存在种内不保守而种间保守的优势表位。分子对接显示实验鉴定的两条优势表位可在人HLAⅡ类超家族分子中均可获得良好的对接分数,进一步肯定其在人群免疫中应用的可行性。结论多肽LFLRATTELRTFSIL和GYYSTTIRYQATGFG可作为EBOV表位疫苗的候选表位。

Objective To analyze the characteristics of pan-MHCⅡrestricted dominant epitopes of Ebola virus glycoprotein and identify candidate epitopes for vaccine development by bioinformatic prediction and downstream analysis.Methods The dominant epitopes of Zaire Ebola virus glycoprotein were predicted by using IEDB-recommended prediction tool and analyzed by cluster analysis,conservation analysis,and molecular docking.Mice were immunized with the plasmid pVAX-GP encoded glycoprotein DNA,and the predicted epitopes were verified by ELISpot assay.Results There were 19 and 13 dominant epitopes obtained from the prediction of restricted 15-mer epitopes binding to the HLAⅡsuperfamily with 99%population coverage and the H-2,respectively.ELISpot assay showed that pVAX-GP induced mouse spleen cells to produce cellular immune responses against synthetic dominant epitopes,among which epitopes LFLRATTELRTFSIL and GYYSTTIRYQATGFG significantly activated cellular immune response in C3H mice.The results of conservation analysis showed that there were dominant epitopes of inter-species conservation and intra-species conservation,yet there were no dominant epitopes of inter-species conservation but no intra-species conservation.Molecular docking showed that the two dominant epitopes identified in the experiment obtained good docking scores in binding to HLAⅡsuperfamily molecules,which further confirmed the feasibility of their application in human immunity.Conclusion The polypeptides LFLRATTELRTFSIL and GYYSTTIRYQATGFG can be used as candidates for the Ebola virus epitope vaccine.