一株抗蓝舌病病毒8型VP2蛋白单克隆抗体识别的线性抗原表位的鉴定

Identification of epitope recognized by a monoclonal antibody against VP2 protein of bluetongue virus serotype 8

摘要:

为研究本实验室制备的一株抗蓝舌病病毒8型 (BTV-8) VP2蛋白的单克隆抗体 (MAb) 3G11识别的B细胞抗原表位，利用噬菌体肽库展示技术对3G11识别的抗原表位进行筛选并鉴定。经过4轮淘选后挑取蓝斑测序，测序结果经分析后获得KLLAT序列，与BTV-8 VP2蛋白氨基酸序列比对后获得共同的短肽序列为283LL284；合成4种短肽序列：KLLAA、KALAT、KLAAT和KLLAT，与3G11细胞上清和腹水分别进行间接ELISA鉴定，结果表明，短肽KLLAA和KLLAT与3G11细胞上清及腹水具有较强的结合能力；与24种BTV标准阳性血清反应结果表明，这两种短肽都可与BTV-8阳性血清发生特异性反应；序列分析结果可见，该表位的氨基酸序列283LL284在不同来源的BTV-8毒株间保守，确定283LL284为MAb3G11识别抗原表位的关键氨基酸。本研究为建立8型BTV特异性的免疫学检测方法和相关病毒蛋白的功能研究奠定了基础。

Abstract:

To confirm the B cell epitope recognized by monoclonal antibody (MAb) 3G11 of bluetongue virus type 8 (BTV-8) VP2 protein prepared in our laboratory, antigen epitopes recognized by 3G11 were screened and identified by phage display technology. KLLAT sequence was found by sequencing of blue spot after four rounds panning and 283LL284 of common short peptide sequence was obtained after comparison to amino acid sequence of BTV-8 VP2 protein. The peptide sequences KLLAA, KALAT, KLAAT and KLLAT were synthesized and identified by indirect ELISA. KLLAA and KLLAT bound strongly with supernatant and as cites of 3G11 cells and reacted specifically with BTV-8 positive standard sera. Further sequence analysis showed that amino acid sequence 283LL284 was conserved among different serotypes of BTV-8 strains, and283LL284 was the key amino acids of antigen epitopes recognized by 3G11. This study laid the foundation to establish type 8 BTV specific immunological detection methods.