阴道毛滴虫硫氧还原蛋白过氧化物酶的cDNA克隆及序列分析

目的获取阴道毛滴虫硫氧还原蛋白过氧化物酶的cDNA克隆,研究其还原烷基氢过氧化物和氢过氧化物的抗氧化作用,以及调节由氢过氧化物介导的信号转换.方法提取阴道毛滴虫的总RNA,用λTripIEx2噬菌体载体构建cDNA表达文库,阳性质粒cDNA克隆进行测序,并用生物软件RPS-Blast,NCBIBlast和ClustalW等对6个读码框架进行序列分析. 结果获得一株开放阅读框为588对碱基的cDNA克隆,推测肽链有196个氨基酸.序列分析表明,该肽链与衣滴虫过氧化物氧化还原酶(Prx1蛋白)及人的自然杀伤增强因子B分别有56%和61%的同源性;除了两个高度保守的半胱氨酸作用基序外,还有蛋白激酶C磷酸化和酪氨酸激酶磷酸化作用基序,N-豆蔻酰化作用位点,脂质运载蛋白信号位点等.结论该蛋白有＞56%的可能性位于胞浆,和典型2-Cys Prx亚家族有很高(56%～62%)的同源性,很可能是其中的一个新成员.

Cloning and characterization of a cDNA encoding thioredoxin peroxidase from Trichomonas vaginalis

Objective:To clone and characterize the thioredoxin peroxidase gene of Trichomonas vaginalis for studying its functions involving not only in the reduction of alkyl hydroperoxide and hydrogen peroxide as an antioxidant, but also in regulating hydrogen peroxide-mediated signal transduction. Methods: T. vaginalis mRNA was isolated to construct a cDNA library using λTripIEx2 phage vector. Sequencing of the positive plasmid cDNAs was performed and the resulted sequences were translated in all six reading frames and compared with the nonredundant databases using the NCBI BlastX, RPS-Blast and ClustalW programs. Results: A cDNA clone was isolated and the sequence analysis revealed that the cDNA clone had an open reading frame with 588 bp. The deduced amino acid sequence contained 196 residues and shared 56% identity with a recently identified Chlamydomonas peroxiredoxin 1 (Prx1) and 61% identity with Human natural killing enhance factor B (NKEFB). The conserved sequence elements including the motif of 2-Cys sites and protein kinase C phosphorylation sites, as well as acid sequence of tyrosine kinase phosphorylation, N-myristoylation and lipocalin signature sites, etc, were detected in the amino acid sequence. Conclusion: The predicted protein located in cytoplasm with more than 56% of reliability presents high identity (56%-62%) with typical 2-Cys Prxs, and it is probably a new member of the typical 2-Cys Prx subfamily.