应用噬菌体肽文库筛选McAb F3特异性结合肽

Study on peptide mimicking epitope of Hantaan virus glycoprotein by phage display peptide library

目的 应用噬菌体展示肽文库筛选可与抗汉坦病毒囊膜蛋白单抗F3特异性结合的配体肽。方法 采用protein A亲和层析纯化的F3单抗为筛选配基 ,对噬菌体展示的随机 12肽文库进行生物亲和淘洗 ;夹心ELISA、竞争ELISA鉴定筛选克隆的结合特性 ,并进一步对阳性克隆进行序列测定和分析。结果 通过 3轮生物淘洗 ,能被抗体捕获的噬菌体克隆为 91.7% (11 12 ) ;ELISA测定显示筛选到的噬菌体短肽能与F3单抗特异性结合 ;序列分析表明 7个阳性克隆氨基酸序列相同 ,均为 MHGPTKNQMWHT ,同源性分析显示该序列与HTNV SEOVM蛋白G2区第 75 0～ 75 9位氨基酸有较高的同源性。结论 获得了具有良好结合活性的模拟表位肽 ,为基于表位水平的HFRSV(肾病综合征出血热病毒 )特异性分子多肽疫苗的设计提供了重要依据。

Objective To obtain the 12-mer phage clones displaying the Hantaan virus mimic epitope related to neutralizing antibody. Methods A group-specific McAb F3 against glycoprotein of both HTNV and SEOV, which possesses fairly high neutralizing titer and has strong animal protective activity. Using F3 McAb as selective molecule, a 12-mer phage peptide library was biopanned for 3 rounds, and the result was confirmed by sandwich ELISA, competition ELISA and DNA sequencing. Results After 3 rounds of effective screening, 11 positive clones screened out from 12 clones were selected, and amino acid sequences of 7 positive clones were consensus: MHGPTKNQMWHT, which was similar to G2 750TKYQYPWHT 759. Conclusion The peptides displayed by phage can mimic the epitope of HFRSV antigen, which provide the potentiality for preparing more effective epitope-based vaccine. [