利用随机肽库鉴定猪瘟病毒E2蛋白抗原表位  

Identification of a Novel Epitope on CSFV E2 Protein by Using Phage Peptide Libraries

作　　者：肖昌[1] 余兴龙[2] 张茂林[2] 徐兴然[2] 涂长春[2] 张玉静[1] 

机构地区：[1]吉林大学畜牧兽医学院 [2]军事医学科学院军事兽医研究所,吉林长春130062

出　　处：《中国兽医学报》2005年第5期449-453,共5页Chinese Journal of Veterinary Science

基　　金：国家"973"计划项目(G1999011903);国家自然科学基金重大项目(39893290-1)

摘　　要：利用噬菌体随机12肽库对抗猪瘟病毒(classical swine fever virus CSFV)糖蛋白E2特异的单抗A11进行表位鉴定,经过4轮筛选后,随机挑取10个噬菌体克隆作竞争ELISA检测.结果表明,10个克隆中除4号克隆外,其余9个均能抑制原核表达的E2蛋白和A11单抗之间的抗原抗体反应,抑制率在35%～64%;DNA测序表明,所有产生竞争抑制作用的8个噬菌体克隆的12肽序列均含有XXWRXXXL核心序列,而没有抑制作用的克隆则不含该核心序列;Western-blot试验证明,所挑阳性克隆均能被单抗A11识别.多序列比较发现,该核心序列与猪瘟病毒E蛋白的28～35位氨基酸TTWKEYSH有一定的同源性,人工合成的含有部分核心序列氨基酸的多肽可以与单抗A11反应,表明单抗A11所针对的抗原表位位于CSFV E2蛋白的28～35位氨基酸.This paper has reported the application of phage peptide libraries to identify a fine antigentic epitope able to react with monoclonal antibody All specifically against immunogenic E2 glycoprotein of classical swine fever virus（CSFV）. After 4 rounds of biopanning using All at gradually decreased concentration a phage peptide library pool was obtained,from which 10 target phage clones were picked up,propagated and tested for their abilities binding to A1 1 McAb in competition with E2 protein in ELISA. The result showed that 9 of 10 those clones could produce 35% to 64% inhibiting efficiency in blocking the reaction of E2 with All ,while one clone showed no competitive effect on E2 binding the McAb. Sequencing of the phage clones resulted in discovery of core peptide sequence XXWRXXXL forming the epitope recognized by All. A specific band with respected molecular weight was visualized in Western blot analysis of those phage clones containing this sequence using McAh All confirming that identification of this epitope was All specific. Alignment of with E2 sequences of pestivirus CSFV,BVDV （bovine viral diarrhea virus） and BDV（border disease virus） pointed A11 core sequence to 28-35 amino acid location of E2 protein although sequence identity is low. Chemically synthesized peptide containing 30 to 42 amino acid region of E2 could react with All further indicating the 28-35 amino acid region is a novel epitope of CSFV.

关 键 词：随机肽库 猪瘟病毒 E2蛋白 抗原表位 

分 类 号：S852.651[农业科学—基础兽医学]