幽门螺杆菌外膜蛋白hopX基因克隆表达及生物信息学分析  

Gene cloning and characterization of hopX outer membrane protein from Helicobacter pylori

作　　者：鞠小丽[1] 邵世和[1] 郝渭滨[1] 

机构地区：[1]江苏大学医学技术学院,镇江212013

出　　处：《中国免疫学杂志》2008年第2期110-113,共4页Chinese Journal of Immunology

基　　金：江苏省科技厅项目(BS2004021)

摘　　要：目的：构建幽门螺杆菌（Helicobacter pylori,Hp）hopX外膜蛋白编码基因的重组质粒,并进行生物信息学分析,探索筛选Hp疫苗的新途径。方法：应用PCR技术从Hp基因组扩增hopX外膜蛋白编码基因片段,TA克隆后测序分析,用信息学软件分析其物理化学特性,并构建表达载体hopX-pQE30转化E.coliM15,IPTG诱导表达,SDS-PAGE及Western blot鉴定表达蛋白。结果：测序分析表明hopX基因全长为1284bp,与Gene Bank公布的其他Hp菌株的基因序列同源性为96%～97%,氨基酸同源性为97%～99%,软件预测显示有多个明显的具有抗原活性的结构域。SDS-PAGE检测表明,47kD处有一新生的蛋白带,Western blot检测重组蛋白具有良好的抗原活性。结论：首次获得HpNCTC11637菌株hopX基因,其表达产物为外膜蛋白生物学功能和疫苗的研究奠定了基础。Objective：To clone the outer membrane protein hopX gene of Helicobacter pylori（Hp）and to perform sequencing and analysis of biological information.Methods：Polymerase chain reaction（PCR）was used to amplify the hopX gene from Hp chromosomal DNA.Then the target gene was digested by restricted endonuclease enzyme of BamH I,and inserted into the prokaryotic expression vector pQE30 digested by corresponding restricted endonuclease enzyme.The recombinant vector was used to select and transform for nucleotide sequence analysis.The biological property at the amino acid level was analyzed by Omiga 2.0 and Antheprot v 5.0.The transformant colony was induced with IPTG and the fusion protein was analyzed by SDS-PAGE and Western blot.Results：The recombinant plasmid was constructed.DNA sequence analysis showed the sequence of hopX was 1 284 bp.The homology of the strains in nucleotide acid was 96%～97%.Their homogeneity in the amino acids was 97%～99%.We get a GeneBank accession number EF208122.Omiga 2.0 software predicted its relative molecular mass（Mr.）was 47 kD and possessed good antigencity.The expressed product contained about 37% of total somatic proteins and Western blot method showed good antigenicity of the recombinant protein.Conclusion：A confirmed gene hopX has been obtained,providing a good foundation for recombination,expression and related study.The corresponding peptide of the gene performed the structural characteristics of some typical antigen molecules,which suggest that it might be a novel vaccine candidate.

关 键 词：幽门螺杆菌 序列分析 生物信息学分析 

分 类 号：R392.1[医药卫生—免疫学]