Macrobrachium rosenbergii cathepsin L: Molecular characterization and gene expression in response to viral and bacterial infections
Macrobrachium rosenbergii cathepsin L: Molecular characterization and gene expression in response to viral and bacterial infections
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DOI：
10.1016/j.micres.2013.04.007
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发表时间：
2013-01-01
影响因子：
6.7
通讯作者：
Kasi, Marimuthu
中科院分区：
文献类型：
--
作者：
Arockiaraj, Jesu;Gnanam, Annie J.;Kasi, Marimuthu
组织蛋白酶L是从构建的罗氏沼虾组织蛋白酶基因文库中鉴定出来的。该基因全长1161个碱基对，开放阅读框架1026个碱基对，编码342个氨基酸的多肽。在该序列的143-154位、286-296位和304-323位分别发现了真核半胱氨酸蛋白酶、组氨酸和天冬酰胺活性中心残基。该基因与日本沼虾组织蛋白L的同源序列相似性最高(97%)，与人类的相似性最低(70%)。系统发育分析显示，在系统发育树上，无脊椎动物和脊椎动物组织蛋白酶L是两个截然不同的类群。日本对虾组织蛋白酶L与日本对虾组织蛋白酶L聚在一起，与斑节对虾组织蛋白酶L形成姊妹群，最后聚为鲑鱼。(P<；0.05)在血细胞中表达水平较高，在眼柄中表达水平最低。此外，罗氏沼气单胞菌在血细胞中的表达受到病毒和白斑综合征杆状病毒(WSBV)1和细菌(哈维氏弧菌和嗜水气单胞菌)的上调。重组蛋白在pH为3~10的范围内表现出较大的活性，在pH为7.5时活性最高。半胱氨酸蛋白酶(Stefin A、Stefin B和止痛剂)对重组的mR-KatL酶活力有显著影响(100%)。对不同浓度的金属离子、盐类和洗涤剂的相对活性和残留活性进行了测试。这些结果表明，金属离子、盐类和洗涤剂对重组菌体蛋白水解酶活性有影响。综上所述，本研究结果表明，组织蛋白酶L具有较高的pH稳定性，是进一步研究组织蛋白酶L在对虾先天免疫系统中作用的研究对象。(C)2013年爱思唯尔股份有限公司。版权所有。
Cathepsin L (MrCathL) was identified from a constructed cDNA library of freshwater prawn Macro-brachium rosenbergii. MrCathL full-length cDNA is 1161 base pairs (bp) with an ORF of 1026 bp which encodes a polypeptide of 342 amino acid (aa) long. The eukaryotic cysteine proteases, histidine and asparagine active site residues were identified in the aa sequence of MrCathL at 143-154, 286-296 and 304-323, respectively. The pair wise clustalW analysis of MrCathL showed the highest similarity (97%) with the homologous cathepsin L from Macrobrachium nipponense and the lowest similarity (70%) from human. Phylogenetic analysis revealed two distinct clusters of the invertebrates and vertebrates cathepsin L in the phylogenetic tree. MrCathL and cathepsin L from M. nipponense were clustered together, formed a sister group to cathepsin L of Penaeus monodon, and finally clustered to Lepeophtheirus salmonis. High level of (P < 0.05) MrCathL gene expression was noticed in haemocyte and lowest in eyestalk. Furthermore, the MrCathL gene expression in M. rosenbergii was up-regulated in haemocyte by virus [M. rosenbergii nodovirus (MrNV) and white spot syndrome baculovirus (WSBV)1 and bacteria (Vibrio harveyi and Aeromonas hydrophila). The recombinant MrCathL exhibited a wide range of activity in various pH between 3 and 10 and highest at pH 7.5. Cysteine proteinase (stefin A, stefin B and antipain) showed significant influence (100%) on recombinant MrCathL enzyme activity. The relative activity and residual activity of recombinant MrCathL against various metal ions or salts and detergent tested at different concentrations. These results indicated that the metal ions, salts and detergent had an influence on the proteinase activity of recombinant MrCathL. Conclusively, the results of this study imply that MrCathL has high pH stability and is fascinating object for further research on the function of cathepsin L in prawn innate immune system. (c) 2013 Elsevier GmbH. All rights reserved.