Molecular characterization and expression analysis of Cathepsin B and L cysteine proteases from rock bream (Oplegnathus fasciatus).
Molecular characterization and expression analysis of Cathepsin B and L cysteine proteases from rock bream (Oplegnathus fasciatus).
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DOI：
10.1016/j.fsi.2010.12.022
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发表时间：
2011-03
影响因子：
4.7
通讯作者：
I. Whang;M. De Zoysa;C. Nikapitiya;Youngdeuk Lee;Yucheol Kim;Sukkyoung Lee;Chulhong Oh;Sung‐Ju Jung;M. Oh;C. Choi;Sang-Yeob Yeo;Bong-Seok Kim;Se-Jae Kim;Jehee Lee
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文献类型：
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作者：
I. Whang;M. De Zoysa;C. Nikapitiya;Youngdeuk Lee;Yucheol Kim;Sukkyoung Lee;Chulhong Oh;Sung‐Ju Jung;M. Oh;C. Choi;Sang-Yeob Yeo;Bong-Seok Kim;Se-Jae Kim;Jehee Lee
组织蛋白酶是木瓜蛋白酶家族的溶酶体半胱氨酸蛋白酶，在溶酶体系统内的细胞内蛋白质降解和翻转中起重要作用。本研究通过对板鱼混合组织cDNA转录组测序，确定了组织蛋白酶B （RbCathepsin B）和组织蛋白酶L （RbCathepsin L）的全长序列。组织蛋白酶B由330个氨基酸残基组成，预测分子量为36 kDa。RbCathepsin L含有336个氨基酸残基，编码一个38 kDa的预测分子质量蛋白。测序分析结果表明，组织蛋白酶B和L均含有木瓜蛋白酶家族的半胱氨酸蛋白酶特征和真核巯基蛋白酶半胱氨酸、天冬酰胺和组氨酸的活性位点。此外，RbCathepsin L含有EF hand Ca2+结合和cathepsin前肽抑制剂结构域。鲷鱼组织蛋白酶B和L与阿根廷Lutjanus argenttimaculatus组织蛋白酶B和Lates calcarifer组织蛋白酶L的氨基酸同源性最高，分别为90%和95%。通过系统发育分析，组织蛋白酶B和L分别与木瓜蛋白酶超家族组织蛋白酶家族成员具有高度的进化关系。实时定量RT-PCR分析结果证实，组织蛋白酶B和L基因在未诱导的石鲷组织中均有表达。此外，在脂多糖（LPS）和迟发爱德华氏菌侵染的肝脏和血细胞中均观察到RbCathepsin B和lmrna的活化，这表明石鲷的免疫应答具有一定的作用。
Cathepsins are lysosomal cysteine proteases of the papain family that play an important role in intracellular protein degradation and turn over within the lysosomal system. In the present study, full-length sequences of cathepsin B (RbCathepsin B) and L (RbCathepsin L) were identified after transcriptome sequencing of rock bream Oplegnathus fasciatus mixed tissue cDNA. Cathepsin B was composed of 330 amino acid residues with 36 kDa predicted molecular mass. RbCathepsin L contained 336 amino acid residues encoding for a 38 kDa predicted molecular mass protein. The sequencing analysis results showed that both cathepsin B and L contain the characteristic papain family cysteine protease signature and active sites for the eukaryotic thiol proteases of cysteine, asparagine and histidine. In addition, RbCathepsin L contained EF hand Ca2+binding and cathepsin propeptide inhibitor domains. The rock bream cathepsin B and L showed the highest amino acid identity of 90 and 95% to Lutjanus argentimaculatus cathepsin B and Lates calcarifer cathepsin L, respectively. By phylogenetic analysis, cathepsin B and L exhibited a high degree of evolutionary relationship to respective cathepsin family members of the papain superfamily. Quantitative real-time RT-PCR analysis results confirmed that the expression of cathepsin B and L genes was constitutive in all examined tissues isolated from un-induced rock bream. Moreover, activation of RbCathepsin B and L mRNA was observed in both lipopolysaccharide (LPS) and Edwardsiella tarda challenged liver and blood cells, indicating a role of immune response in rock bream.