Macrobrachium rosenbergii cathepsin L: molecular characterization and gene expression in response to viral and bacterial infections.
Cathepsin L (MrCathL) was identified from a constructed cDNA library of freshwater prawn Macrobrachium rosenbergii. MrCathL full-length cDNA is 1161 base pairs (bp) with an ORF of 1026bp 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)] 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.
['Arockiaraj|Jesu|J|', 'Gnanam|Annie J|AJ|', 'Muthukrishnan|Dhanaraj|D|', 'Thirumalai|Muthukumaresan Kuppusamy|MK|', 'Pasupuleti|Mukesh|M|', 'Milton|James|J|', 'Kasi|Marimuthu|M|']
D000595:Amino Acid Sequence D000818:Animals D001419:Bacteria D056668:Cathepsin L D016000:Cluster Analysis D018076:DNA, Complementary D004791:Enzyme Inhibitors D004795:Enzyme Stability D020869:Gene Expression Profiling D006863:Hydrogen-Ion Concentration D008969:Molecular Sequence Data D016366:Open Reading Frames D033541:Palaemonidae D010802:Phylogeny D011994:Recombinant Proteins D017422:Sequence Analysis, DNA D017386:Sequence Homology, Amino Acid D014780:Viruses
Cathepsin L Nodovirus Baculovirus Bacteria Enzyme activity Machrobrachium rosenbergii