摘要： 对溶藻弧菌外膜蛋白OmpU进行分子克隆构建,抗原性生物信息学分析,为疫苗的开发奠定了基础.通过分子克隆获得OmpU蛋白的原核表达菌株,重组表达后利用切胶纯化获得OmpU蛋白,免疫小鼠制备抗血清,Western Blotting检测发现OmpU蛋白具有很好的抗原性.采用在线软件预测OmpU为亲水的分泌型蛋白;存在多个酶切位点.采用TMHMM Server v.2.0程序预测OmpU无跨膜结构并定位于细胞膜外.通过SignalP 4.1软件分析发现OmpU的1~21位氨基酸为信号肽序列.SOPMA服务器预测OmpU的二级结构含无规则卷曲27.94%,α-螺旋为33.24%,β-转角为10.29%,β-片层为28.53%.采用Swiss Model程序预测三维结构显示OmpU具有3个孔道结构.综合利用ABCpred和BepiPred方案,预测OmpU存在8个B细胞表位.运用神经网络法预测OmpU具有3个CTL表位.使用MHC-Ⅱ类分子结合肽在线程序预测OmpU存在1个Th表位.

关键词: 溶藻弧菌, OmpU蛋白, 细胞表位, 蛋白结构, 预测

Abstract: The construction of molecular cloning,antigenicity identification and bioinformatics analysis of Vibrio alginolyticus outer membrance protein U(OmpU),laid the foundation for the development vaccine.The OmpU expression strain was obtained by molecular clone.OmpU was purified by the way of the gel slices,mice immunized were to prepare antiserum and found that OmpU had better antigenicity using Western Blotting method.By online software,the prediction results showed that OmpU protein was a hydrophilic and secreted protein multiple restriction enzyme sites.OmpU secondary structure contained random coil 27.94%,alpha helix 33.24%,beta turn 10.29%,β sheet 28.53% by SOPMA method.Using TMHMM Server v.2.0 software predicted that OmpU had not transmembrane structure and was located outside cell membrane,and the SignalP 4.1 software found that the 1-21 amino acids of OmpU was signal peptide sequence.The 3D structure prediction found that OmpU protein had 3 core structure.Using ABCpred and BepiPred prediction program,OmpU had 8 B cell epitopes.By the prediction method of artificial neural network,OmpU protein had 3 CTL epitopes.OmpU protein had 1 Th epitopes using an online server prediction for MHC class Ⅱ peptide binding affinity.

Key words: Vibrio alginolyticus, OmpU protein, Cell epitopes, Protein structure, Prediction

中图分类号: 

• S942

• Q78