布鲁氏菌S2株L7/L12蛋白B细胞线性抗原表位的预测与鉴定  

Prediction and preliminary evaluation of B-cell linear epitopes in the L7/L12 protein of Brucella suis S2

作　　者：成岩[1,2] 白靓[3] 董丽刚 韩梅[1,2] 王敏[1,2] 张冬丽[1,2] 霍万学[1,2] 

机构地区：[1]内蒙古民族大学医学院,内蒙古通辽028000 [2]内蒙古民族大学病原生物学与免疫学研究所 [3]内蒙古民族大学生命科学学院

出　　处：《中国病原生物学杂志》2015年第3期206-210,共5页Journal of Pathogen Biology

基　　金：内蒙古自治区自然科学基金项目(No.2011MS1107)

摘　　要：目的预测猪布鲁氏菌S2株L7/L12蛋白B细胞线性抗原表位,并鉴定其抗原性。方法以布鲁氏菌S2株为对象,采用NCBI提供的BLAST程序分析其L7/L12蛋白与原核模式生物大肠埃希菌L7/L12蛋白的同源性;使用IEDB网络服务器初步预测B细胞线性表位;在大肠埃希菌L7/L12蛋白空间结构的基础上,使用SWISS-MODEL服务器构建猪布鲁氏菌S2株L7/L12蛋白空间结构模型,参考空间结构特点筛选符合形成表位的区域,同时分析区域内易形成表位的3种氨基酸(缬氨酸、亮氨酸、半胱氨酸)所在位置,综合分析以上结果获得所预测表位;以预测表位序列为基础,人工合成表位多肽-匙孔血蓝蛋白复合抗原并包被酶标板,分别以rL7/L12多克隆抗体及小鼠布鲁氏菌免疫血清为一抗,采用间接ELISA方法验证所预测表位的抗原活性。结果 L7/L12含有潜在的B细胞线性抗原表位,所预测表位区位于N末端第51-66aa,60-75aa,87-102aa;ELISA检测合成的表位多肽蛋白复合物抗原性,60-75aa表位能够与rL7/L12多克隆抗体及布鲁氏菌免疫鼠血清抗体结合。结论筛选的猪布鲁氏菌L7/L12蛋白B细胞线性表位位于N末端第60-75aa区域,具有与特异性血清结合的能力,为布鲁氏菌的感染诊断与疫苗研发奠定了基础。Objectives To predict B-cell linear epitopes in the L7/L12 protein of Brucella suis S2 and to evaluate the antigenicity of the predicted epitopes. Methods The program BLAST from the NCBI was used to analyze the similarity of the L7/L12 protein of B.suis S2 and the L7/L12 protein of E.coli.A server networked to the IEDB was used to preliminarily predict B-cell linear epitopes.Based on the spatial structure of the L7/L12 protein of E.coli,the spatial structure of the L7/L12 protein of B.suis S2 was modeled using SWISS-MODEL.In light of the characteristics of the spatial structure,the protein was screened for corresponding regions containing epitopes.The location of three amino acids（valine,leucine,and cysteine）was also analyzed and identified in the epitopes.Epitopes were verified through comprehensive analysis of the results of those analyses.In order to predict B-cell linear epitopes,apeptide-keyhole limpet hemocyanin conjugate was prepared as a coating antigen,and rL7/L12 polyclonal antibodies and mouse serum that was positive for Brucella were used as the primary antibodies.Indirect ELISA was used to verify the antigenicity of the predicted epitopes.Results Potential B-cell linear epitopes in L7/L12 were located at amino acids（aa）51-66,60-75,and 87-102 of the N-terminal region.Indirect ELISA results indicated that the peptide-protein conjugate is antigenic and can bind to rL7/L12 polyclonal antibodies and mouse serum that was positive for Brucella. Conclusion Screening indicated that the L7/L12 protein had a B-cell linear epitope at aa 60-75 of the N-terminal region.Serum antibodies can specifically bind to this epitope.This study has laid the foundation for research into the diagnosis of brucellosis and vaccines against Brucella.

关 键 词：布鲁氏菌 L7/L12 表位 生物信息学 

分 类 号：R378.5[医药卫生—病原生物学]