利用酵母随机展示文库筛选与鉴定基因Ⅶ型新城疫病毒HN蛋白B细胞抗原表位

Screening and identification of a B-cell epitope on HN protein of genotype Ⅶ Newcastle disease virus using yeast random display library

新城疫病毒(Newcastle disease virus,NDV)给全世界家禽养殖业造成了严重的经济损失。HN蛋白是NDV上的重要囊膜蛋白,能诱导鸡体内的免疫系统产生保护性抗体。为了鉴定HN蛋白上的B细胞表位,本研究成功构建了基因Ⅶ型NDV HN蛋白的酵母随机展示文库,该文库的库容量为5×10^5 CFU,插入的片段随机分布。利用NDV高免血清标记HN蛋白的酵母随机展示文库,然后通过流式细胞分选技术分选阳性克隆细胞进行测序分析。结果显示,阳性文库中150号多肽(GAPVHDPDYIGGIGKE)含量很高。对150号多肽进行截短分析,结果表明该多肽中VHDPDYIGGI序列为最小的B细胞表位,该表位能与感染NDV的高免血清普遍反应。因此,VHDPDYIGGI多肽是基因Ⅶ型NDV HN蛋白中广泛存在的B细胞表位。本研究结果为将来开发新城疫新型ELISA检测试剂盒以及表位疫苗奠定了基础。

Newcastle disease virus(NDV)has caused severe economic losses to the poultry industry around the world.HN protein is an important envelope protein on NDV and can induce the production of protective antibodies in chickens.To identify the B cell epitope in HN protein,a yeast random display library of HN protein of genotypeⅦNDV was successfully constructed in this research.The library size was 5×10^5 CFU,and the inserts were randomly distributed.The positive cells in the yeast library were screened with flow cytometry after being stained with high immune serum of NDV and sequenced for the analysis of the inserts.The results showed that the peptide 150(GAPVHDPDYIGGIGKE)was highly detected in the positive library.The truncation analysis showed that the B cell epitope in the peptide 150 was VHDPDYIGGI,and this epitope was able to react with the high immune sera from chickens infected with NDV.Therefore,the identified peptide(VHDPDYIGGI)is a general B-cell epitope presented in the HN protein of NDV.These results provide the foundation for the development of a new ELISA detection kits and epitope vaccines in the future.