八肋游仆虫组织蛋白酶B基因家族的分子克隆及序列分析

Molecular Cloning and Sequences Analysis of Cathepsin B Gene Family in the Ciliate(Euplotes octocarinatus)

组织蛋白酶B (cathepsin B, CTSB)是一种主要存在于溶酶体中的半胱氨酸蛋白水解酶,广泛存在于各种生物中。为了研究八肋游仆虫组织蛋白酶B (Euplotes octocarinatus Cathepsin B, EoCTSB)基因的序列、结构和功能,本研究利用生物信息学方法,对八肋游仆虫组织蛋白酶B基因进行了系统分析。利用相似性搜索及系统进化分析,共鉴定得到6个EoCTSBs基因。转录组数据及3′RACE分析结果表明这6个基因均具有转录活性。通过与其他真核生物的组织蛋白酶B比较,结果显示,6种EoCTSBs均含有保守的肽链内切酶催化活性位点,但都缺失封闭环结构,因此推测它们都没有外肽酶活性;仅EoCTSB-6的S2亚位为酸性氨基酸残基,暗示其具有催化特异性底物Z-Arg-Arg-AMC水解的活性。本研究为后续深入探讨EoCTSB的生物学功能提供理论依据。

Cathepsin B(CTSB) is a cysteine protease that is mainly found in lysosomes and widespread in a variety of organisms. To study the sequence, structure and function of the Cathepsin B from Euplotes octocarinatus(named as EoCTSB), we systematically analyzed the Cathepsin B gene of Euplotes octocarinatus by using bioinformatics methods. Using similarity search and phylogenetic analysis, 6 EoCTSB genes were identified.Transcriptional activity of EoCTSB genes were verified by transcriptome data and 3’RACE analysis. Comparison of EoCTSBs with other eukaryotes CTSBs showed that all six EoCTSBs contain conserved endopeptidase catalytic sites, but all of them lack the occluding loop structure. Therefore, we speculate that none of them have exopeptidase activity. Only the S2 subsite of EoCTSB-6 is an acidic amino acid residue, suggesting that it has the activity of catalyzing the hydrolysis of the specific substrate Z-Arg-Arg-AMC. This study provides a theoretical basis for further exploration of the biological functions of EoCTSB.