细粒棘球绦虫Calpain蛋白的生物信息学分析

Bioinformatic analysis of calpain from Echinococcus granulosus

目的采用生物信息学软件分析细粒棘球绦虫Calpain蛋白(EgCalpain)的结构与抗原表位,为研制基于Calpain的棘球蚴病疫苗提供依据。方法从NCBI核苷酸和蛋白质数据库下载EgCalpain的核苷酸和氨基酸序列,采用ProtParam、SignalP、TMHMM、Cell-Ploc等在线分析程序,结合DNAStar、DNAMAN等生物信息学软件分析、预测EgCalpain蛋白的理化性质、信号肽序列、跨膜结构域、亚细胞定位及抗原表位;采用Clustal omega程序对不同物种来源的Calpain蛋白进行多序列比对,并通过MEGA 6.06程序构建邻接树。结果 EgCalpain基因全长7 734 bp,包含15个外显子和14个内含子;EgCalpain基因编码蛋白由707个氨基酸组成,是一种稳定蛋白,不含有信号肽序列和跨膜区域,可能定位于细胞质。EgCalpain蛋白含有5个T、B细胞联合表位,分别为267-299 aa、328-356 aa、486-507 aa、546-567 aa、648-662 aa。EgCalpain蛋白与曼氏血吸虫Calpain的亲缘关系较近,与人、牛、羊Calpain的亲缘关系较远。结论 EgCalpain蛋白含有5个T、B细胞联合表位且与人、牛、羊Calpain的亲缘关系较远,作为棘球蚴病疫苗进行免疫时,可能具有良好的免疫保护作用。

Objective To analyze the structure and epitopes of calpain from Echinococcus granulosus(EgCalpain) with bioinformatic software in order to lay the foundation for the production of echinococcosis vaccines based on calpain. Methods Nucleotide and amino acid sequences of EgCalpain were downloaded from nucleotide and protein databases on the NCBI website. Bioinformatic tools, such as ProtParam, SignalP, TMHMM, and Cell-Ploc, were used to analyze physiological and biochemical characteristics, signal peptide sequence, transmembrane domains, and sub-cellular localization. T-and B-cell epitopes were predicted using DNAStar and DNAMAN. Multiple sequence alignment of calpain from various species was performed using Clustal omega, and a neighbor-joining tree was constructed using MEGA 6.06. Results EgCalpain gene was 7 734 bp in length, and it had 15 exons and 14 introns. The protein coded for by the EgCalpain gene was a stable protein consisting of 707 amino acids and located in the cytoplasm. No signal peptide sequences or transmembrane domains were found in the amino acid sequences of EgCalpain. EgCalpain protein was predicted to have 5 combined T-and B-cell epitopes that were located at amino acids 267-299, 328-356, 486-507, 546-567, and 648-662. EgCalpain was evolutionarily close to calpain from Schistosoma mansoni(SmCalpain) but distant from calpain from humans, Bos taurus, and Ovis aries. Conclusion EgCalpain protein contained five combined T-and B-cell epitopes, and it was evolutionarily distant from calpain from humans, Bos taurus, and Ovis aries. All of these findings indicated that an EgCalpain vaccine can provide good immune protection.