Molecular cloning and functional characterization of a novel i‐type lysozyme in the freshwater mussel Cristaria plicata

Abstract

The freshwater bivalve Cristaria plicata, which is widely distributed in Eastern Asia, is a key species in the pearl culture industry. In this study, a novel invertebrate-type lysozyme, designated as CpLYZ2, was cloned from hemocytes of C. plicata. This lysozyme shares high sequence identity and is homologous to a previously identified lysozyme CpLYZ1 isolated from C. plicata and with HcLyso3 isolated from Hyriopsis cumingii. The full-length cDNA of CpLYZ2 is 913 bp long, which includes an open reading frame (ORF) of 486 bp, a 3 0 untranslated region (UTR) of 389 bp and a 5 0 UTR of 38 bp. The ORF encodes a putative polypeptide of 161 amino acids with a predicted molecular mass of 18.2 kDa and a theoretical isoelectric point of 6.56. CpLYZ2 mRNA transcripts can be detected in hemocytes, hepatopancreas, muscle, gills and mantle tissues, the greatest expression being observed in the gills. CpLYZ2 expression in hemocytes, hepatopancreas and gills increased significantly after the mussel was challenged with Aeromonas hydrophila. Furthermore, the optimal pH and temperature for enzyme activity of the recombinant CpLYZ2 were 5.5 and 50°C, respectively. The recombinant lysozyme protein exhibited bacteriolytic activity against Escherichia coli, A. hydrophila, Staphyloccocus aureus, Bacillus subtilis, Streptococcus sp. and Staphylococcus epidermidis. The findings of this study help to elucidate immune responses in molluscs and will thus expedite disease management of these key freshwater species, in turn boosting pearl culture in eastern Asia.

Seasonal Changes in Hemolymph Parameters of the Bivalve Modiolus kurilensis Bernard, 1983 from Vostok Bay, Sea of Japan

“…These results suggest the presence of lytic factors in the 16 kDa fraction. Moreover, the range of molecular mass of lytic factors (especially lysozymes) varies from 12 to 22 kDa for many mollusks: M. galloprovincialis (C-type lysozyme 14.4 kDa), R. philippinarum (invertebrate type lysozyme 20.87 kDa), Crassostrea virginica (i-type lysozymes 12.98 and 17.86 kDa), Cristaria plicata (i-type lysozyme 18.2 kDa), Meretrix meretrix (i-type lysozyme 14.6 kDa) [39][40][41][42][43][44].…”

Applications of Lysozyme, an Innate Immune Defense Factor, as an Alternative Antibiotic

“…The lysozyme from the viscera of scallops ( Patinopecten yessoensis ) was purified and characterized in 2008 [ 40 ]. With the same aim of identifying new, more active enzymes, the lysozymes of other bivalve mollusks, including Unio pictorum [ 41 ] (four-fold more active than the egg lysozyme in the inhibition of E. coli ); Asian hard clams ( Meretrix meretrix ) [ 42 ]; freshwater mussels ( Cristaria plicata ) [ 43 ]; and Manila clams ( Ruditapes philippinarum ) [ 44 ], were isolated and identified.…”

Exploring the therapeutic potential of recombinant human lysozyme: a review on wound management system with antibacterial

“…Bacillus subtilis is another attractive host due to its Generally Recognized As Safe (GRAS) status, secretion capabilities, and lack of endotoxins. Engineered B. subtilis strains with increased protease deficiency and simplified downstream processing have achieved rhLZM titers of approximately 500 mg/ L (Dai et al, 2015). Yeast systems, such as Pichia pastoris and Saccharomyces cerevisiae, are also gaining traction for secreting rhLZM with yields exceeding 1 g/L (Symmank et al, 2002).…”