旋毛虫硫氧还蛋白过氧化物酶基因的克隆与生物信息学分析

Cloning and bioinformatics analysis of thioredoxin peroxidase genes fromTrichinella spiralis

为了研究旋毛虫硫氧还蛋白过氧化物酶的功能,对GenBank中旋毛虫基因组数据库和旋毛虫EST数据库进行检索,获得旋毛虫硫氧还蛋白过氧化物酶基因的cDNA序列并设计引物。以旋毛虫肌幼虫总RNA为模板,进行RT-PCR,将PCR产物克隆到pMD18-T载体后测序并进行生物信息学分析。结果显示,成功克隆到3个旋毛虫硫氧还蛋白过氧化物酶基因的cDNA序列(TsTPx1,TsTPx2,TsTPx3)。TsTPx1cDNA含有1个由747个核苷酸组成的完整的开放阅读框,编码248个氨基酸残基组成的多肽,蛋白质的理论分子质量为28.2ku,理论等电点为9.00。TsTPx2cDNA含有1个由588个核苷酸组成的完整的开放阅读框,编码195个氨基酸残基组成的多肽,蛋白质理论分子质量为22.1ku,理论等电点为6.52。TsTPx3cDNA含有1个由594个核苷酸组成的完整的开放阅读框,编码197个氨基酸残基组成的多肽,蛋白质理论分子质量为22.4ku,理论等电点为6.08。TsTPx1、TsTPx2、TsTPx3都具有1个AhpC-TSA结构域和1个1-cys Prx_C结构域。同源性分析表明,TsTPx1、TsTPx2、TsTPx3与其他线虫硫氧还蛋白过氧化物酶的一致性在60%左右。

In order to study the function of thioredoxin peroxidase of Trichinellaspiralis,primers derived fromT.spiralis genome and EST database in GenBank were designed and the cDNA sequence of thioredoxin peroxidase genes(TsTPx1,TsTPx2and TsTPx3)were cloned by RT-PCR fromT.spiralis muscle larvae and bioinformatics analysis was performed.In result,the TsTPx1cDNA sequence contained an open reading frame(ORF)of 747nucleotides and the deduced protein consisted of 248amino acids with the theoretical molecular weight of 28.2ku and an isoelectric point of 9.00.TsTPx2cDNA sequence contained an ORF of 588nucleotides and the deduced protein consisted of 195amino acids with a theoretical molecular weight of 22.1ku and isoelectric point of 6.52.TsTPx3cDNA sequence contained an ORF of 594nucleotides and the deduced protein consisted of 197amino acids with a theoretical molecular weight of 22.4ku and an isoelectric point of 6.08.All the deduced proteins had no signal peptide sequence,while they all had an AhpC-TSA domain and a 1-cysPrx_C domain.Amino acid sequences of TsTPx1,TsTPx2,and TsTPx3 shared about 60%identity with those from other parasitic nematodes.