猪乙型脑炎病毒广西分离株FC792的囊膜糖蛋白E基因克隆及生物信息学分析  

Cloning and Bioinformatics Analysis of Envelope Glycoprotein E Gene of Swine Japanese Encephalitis Virus Strain FC792 Isolated from Guangxi

作　　者：李斌[1,2] 卢冰霞[1,2,3] 秦毅斌[1,2] 赵武[1,2] 梁家幸[1,2] 韦超文 何颖[1,2] 苏乾莲[1,2] 陈忠伟[1,2] 闭炳芬[1,2] 段群棚[1,2] 李莹莹[1,2,3] 姜佳佳[1,2,3] 梁保忠[1,2] 黄伟坚[3] 

机构地区：[1]广西兽医研究所,南宁530001 [2]广西畜禽疫苗新技术重点实验室,南宁530001 [3]广西大学动物科学技术学院,南宁530005 [4]广西田林县动物疫病预防控制中心,田林533300

出　　处：《基因组学与应用生物学》2013年第4期467-478,共12页Genomics and Applied Biology

基　　金：广西自然科学基金项目(2011GXNSFB018032);公益性行业(农业)科研专项(201103008);广西科技攻关项目(桂科攻1123007-3);广西畜禽疫苗新技术重点实验室系统性研究课题(12-071-28-A-5);广西基本科研业务费专项桂科(专项13-1)共同资助

摘　　要：为了研究开发快速、敏感、特异的猪乙型脑炎(Japanese encephalitis,JE)诊断试剂和免疫效果良好的猪JE基因工程疫苗,对2011年分离到的广西猪乙型脑炎病毒(Japanese encephalitis virus,JEV)FC792株的E基因进行克隆、测序及生物信息学分析。结果测序获得1500bp的完整FC792株E基因序列,经同源性比对及遗传进化树分析表明,FC792株为基因Ⅲ型JEV毒株,且属于弱毒株。应用在线软件SignalP 4.1对FC792株E蛋白进行信号肽的预测,结果预测到FC792株E蛋白不存在信号肽。应用在线软件NetNGlyc 1.0预测到FC792株E蛋白存在1个N-糖基化位点:154-NYSA,线软件NetPhos 2.0预测到其存在如下磷酸化位点:丝氨酸(Ser)有10个位点;苏氨酸(Thr)有6个位点;酪氨酸(Tyr)有5个位点。应用在线免疫信息学软件预测到FC792株E蛋白有16个B细胞抗原优势表位(分值(score)≥0.85),5个CTL抗原表位,10个Th抗原表位。应用在线软件GOR4预测FC792株E蛋白的二级结构,发现α-螺旋区域占21.20%,延伸链区域占30.40%;无规则卷曲区域占48.40%。应用在线软件SWISS-MODEL对FC792株E蛋白进行三维结构预测的同源模建,结果可见FC792株E蛋白存在大量的无规则卷曲和延伸链区域,并有多个α-螺旋区域,它们螺旋扭转卷曲形成近似左右对称的2个亚基的二聚体空间立体结构。研究结果为今后研制猪JE诊断抗原和猪JE基因工程疫苗提供参考借鉴。In order to study and develop a rapid, sensitive and specific Japanese encephalitis （JE） diagnostic reagents and adequate immune response to swine JE genetically engineered vaccine, we fully analyzed E gene clone, se- quence and bioinformatics analysis study of a so-called FC792, Guangxi swine Japanese encephalitis virus （JEV）strain isolated in year 2011. As results, we obtained a 1 500 bp full length of FC792 strains＇ E gene sequence; through homology comparison and phylogenetic analysis, the isolate FC792 belongs to attenuated JEV genotype ]]1. Using online software SignalP 4.1 to forecast FC792 strain＇s E protein signal peptide, results showed absence of E protein signal peptide. Using online software NetNGlyc 1.0, we could forecast the existence of one N-glucosylation site： 154-NYSA, NtPhos 2.0 could forecast the existence of the following phosphorylation sites： Serine （Ser） with 10 sites, Threonine （Thr） with 6 sites, Tyrosine （Tyr） with 5 sites. Application of online immunoinformatics analy- sis, we predetermined that FC792 had 16 B cells epitopes antigens （score 1〉 0.85）, 5 CTR epitopes, 10 Th epitopes. While analysis with online software GOR4 forecasting of FC792 strain, E protein＇s secondary structure, we found an a-helix region to stand by 21.20%, the extended strand region 30.40%, the random coil region 48.40%. Online software SWISS-MODEL processing FC792 strain＇s E protein three dimensional structure＇s homology modeling resulted in the existence of an abundant random coil and extended strand regions and equally a number of a-helix region; their spiral twist curly shapes in two symmetric subunit dimer stereochemical structure. These finding pro- vided references for a henceforth development of Swine JE diagnostic kit and genetically engineered vaccine.

关 键 词：猪 乙型脑炎病毒 E基因 克隆 生物信息学 

分 类 号：S852.65[农业科学—基础兽医学]