日本血吸虫组织蛋白酶L家族1个新基因的克隆与序列分析  

Clone and sequence analysis of a new schistosoma japonicum cathepsin L family protease gene

作　　者：陈奕慧[1] 李孜[2] 

机构地区：[1]广州医学院第二附属医院医院感染管理科,广东广州510260 [2]广州医学院病原生物学教研室,广东广州510182

出　　处：《中华医院感染学杂志》2011年第12期2404-2407,共4页Chinese Journal of Nosocomiology

基　　金：国家自然科学基金(No30600516);广东省自然科学基金(No5300973)

摘　　要：目的克隆与分析日本血吸虫组织蛋白酶L样(SjCLs)基因全长cDNA序列,为进一步研究其功能奠定基础。方法提取日本血吸虫成虫总RNA,经RT-PCR扩增获得cDNA序列;将目的片段连接至pMD18-T Simple载体上,连接产物转化大肠埃希菌,通过酶切和测序鉴定重组质粒;利用生物信息学软件分析序列的开放阅读框、蛋白序列的同源率和功能区。结果以成虫总RNA为模板进行RT-PCR,得到一段长为996 bp的cDNA片段,该片段含有完整读码框,该阅读框编码的蛋白含有331个氨基酸残基,分子量为38.3 kDa,命名为SjCLs;序列比对分析发现该蛋白氨基酸序列与SjCL2同源率最高,达到89.1%;进化树分析发现SjCLs与SjCL2亲缘关系也最近;SjCLs具有类似于SjCL2的N端信号肽,可能经加工后分泌到细胞外;两者所预测的功能基序及三维构象也基本相同。结论 SjCLs具有组织蛋白酶L的基本特征,从而证明研究获得组织蛋白酶L基因家族的一个新成员。OBJECTIVE To clone and analyze the cDNA of Schistosoma japonicum cathepsin L-like protease gene（SjCLs）,which lays a foundation for further investigation of the physiological function of SjCLs.METHODS Total RNA was extracted from adult Schistosoma japonicum.cDNA was obtained from RT-PCR and then cloned into the pMD18-T Simple vector.The recombinant construct was confirmed by restriction endonuclease digestion and DNA sequencing.The open reading frame（ORF） of the SjCLs cDNA sequence,the amino acid identity and the functional regions of protein sequence were analyzed by bioinformatics software.RESULTS A 996 bp cDNA fragment was obtained by RT-PCR.The longest ORF in the cDNA sequence encodes a putative protein of 331 amino acids.The putative protein with molecular weight of 38.3 kDa was named as SjCLs.The amino acid identity between SjCLs and SjCL2 was up to 89.1%.Phylogenetic analysis showed that SjCLs has the closest genetic relationship with SjCL2,comparing to other cathepsins.A Nterminal signal peptide,which was similar to that of SjCL2,was predicted on SjCLs,implicating that SjLs might be processed and then sorted into extracellular.The putative functional domains and the three-dimensional conformation of SjCLs were similar to those of SjCL2.CONCLUSION Sequence analysis shows that SjCLs shares the same characteristics with cathepsin L,suggesting that a new member of the cathepsin L family has been obtained in this study.

关 键 词：日本血吸虫 组织蛋白酶 基因 克隆 序列 分析 

分 类 号：R383.24[医药卫生—医学寄生虫学]