应用噬菌体肽文库筛选肾综合征出血热病毒优势B细胞结合表位肽  

Selection of HFRSV-specific epitopes by screening random peptide libraries with HFRSV-positive sera

作　　者：王倩[1] 俞守义[2] 王长军 

机构地区：[1]南方医科大学南京临床医学院、南京军区南京总医院医务部,江苏南京210002 [2]南方医科大学流行病学教研室,广东广州510515 [3]南京军区疾病控制中心,江苏南京210002

出　　处：《医学研究生学报》2008年第5期475-479,共5页Journal of Medical Postgraduates

摘　　要：目的：应用噬菌体展示肽文库筛选鉴定肾综合征出血热病毒优势B细胞结合表位肽。方法：以Prote in-A纯化的肾综合征出血热患者恢复期血清为筛选配基,对经正常人血清预吸附的噬菌体展示随机12肽文库（或次级肽文库）进行生物亲和淘选;夹心ELISA、竞争ELISA鉴定所获得克隆的结合特性,并进一步对阳性克隆进行序列测定和分析。结果：ELISA测定显示,通过5轮淘选得到的噬菌体克隆半数以上可与筛选血清发生特异性结合;对45个阳性克隆进行序列测定分析,根据其推导的氨基酸序列可大致分为8组,同源性分析显示,Ⅰ-Ⅶ组序列基序可能为LVXKR、LTXR、IXKP、LXPA、VGA、KX IR、EKXP,其中4组基序在病毒结构蛋白氨基酸序列中发现同源区。结论：获得了一组肾综合征出血热病毒（HFRSV）优势B细胞表位肽,HFRSV核蛋白可能是诱导高水平抗体的主要免疫原,研究结果为基于表位的基因工程诊断试剂的研制和多肽疫苗的设计提供依据。Objective： To obtain the 12-mer phage clones displaying the Hantaan virus mimic epitopes. Methods： HFRSV-positive sera were used as selective molecules for biopanning. A 12-mer phage peptide library was bio-panned for 5 rounds, and the result was confirmed by sandwich ELISA, competition ELISA and DNA sequencing. Results： After 5 rounds of effective screening, the results detected by sandwich ELISA and competition ELISA showed that the majority of the selected clones could react to positive sera in a dose-dependent manner, but could not bind to BSA and the control sera. Sequencing and alignment analyses indicated that amino acid sequences of 45 positive clones fell into 8 groups, and 7 of them exhibited putative motifs ： LVXKR, LTXR, IXKP, LXPA, VGA, KXIR and EKXP. Four of the putative motifs had a homologous region within the structural proteins of HFRSV.Conclusion ： The peptides displayed by the phage can mimic the epitopes of HFRSV antigens, which provides the potential for preparing more effective epitope-based vaccines and specific diagnostic reagents.

关 键 词：肾综合征出血热病毒 噬菌体展示肽文库 抗体 表位 

分 类 号：Q78[生物学—分子生物学]