Analysis of Two Lysozyme Genes and Antimicrobial Functions of Their Recombinant Proteins in Asian Seabass

Abstract

Lysozymes are important proteins of the innate immune system for the defense against bacterial infection. We cloned and analyzed chicken-type (c-type) and goose-type (g-type) lysozymes from Asian seabass (Lates calcarifer). The deduced amino acid sequence of the c-type lysozyme contained 144 residues and possessed typical structure residues, conserved catalytic residues (Glu50 and Asp67) and a “GSTDYGIFQINS” motif. The deduced g-type lysozyme contained 187 residues and possessed a goose egg white lysozyme (GEWL) domain containing three conserved catalytic residues (Glu71, Asp84, Asp95) essential for catalytic activity. Real time quantitative PCR (qRT-PCR) revealed that the two lysozyme genes were constitutively expressed in all the examined tissues. The c-type lysozyme was most abundant in liver, while the g-type lysozyme was predominantly expressed in intestine and weakly expressed in muscle. The c-type and g-type transcripts were up-regulated in the kidney, spleen and liver in response to a challenge with Vibrio harveyi. The up-regulation of the c-type lysozyme was much stronger than that of the g-type lysozyme in kidney and spleen. The recombinant proteins of the c-type and g-type lysozymes showed lytic activities against the bacterial pathogens Vibrio harveyi and Photobacterium damselae in a dosage-dependent manner. We identified single nucleotide polymorphisms (SNPs) in the two lysozyme genes. There were significant associations of these polymorphisms with resistance to the big belly disease. These results suggest that the c- and g-type genes play an important role in resistance to bacterial pathogens in fish. The SNP markers in the two genes associated with the resistance to bacterial pathogens may facilitate the selection of Asian seabass resistant to bacterial diseases.

Trial on using of some herbal extracts as promising immunoprophylaxis feed additives in cultured Oreochromis niloticus

“…Lysozyme activity provides an insight into the immune response of fish [32,41]. Results indicated an increase in serum lysozyme activity of both fish groups fed on onion and barley extracts.…”

Molecular characterisation, evolution and expression analysis of g-type lysozymes in Ciona intestinalis

“…In particular, in CiLys-g3 all three catalytic residues (Glu, Asp, Asp) responsible for the binding of the lysozyme to the bacterial cell wall turned out to be highly conserved (Fu et al, 2013) whereas in CiLys-g1 and CiLys-g4, only two catalytic residues (Glu, Asp) were found to be conserved. Structure and surface analysis revealed that the conserved GXXQ motif among the Cilys-g lysozymes is located nearer to the hydrophobic core region.…”

Molecular characterization and expression pattern of c-type and g-type lysozyme isoforms in starry flounder Platichthys stellate infected with Streptococcus parauberis

“…Since the starry flounder c-type lysozyme contains aspartic acid residues at positions 100 and 101 that have been known to be necessary for calcium binding, it may belong to the calcium binding family, likely with mammalian and avian orthologues [30][31][32]. G-type lysozyme in starry flounder possesses conserved residues essential for catalytic activity, which would be commonly observed in orthologues from various animal species [2,5,8]. In the alignment, Glu 72 and Asp 96 (starry flounder numbering) are conserved in all species, whereas the Asp 85 residue is present in all teleostean sequences except common carp Cyprinus carpio [5,8,[33][34][35].…”

“…G-type lysozyme in starry flounder possesses conserved residues essential for catalytic activity, which would be commonly observed in orthologues from various animal species [2,5,8]. In the alignment, Glu 72 and Asp 96 (starry flounder numbering) are conserved in all species, whereas the Asp 85 residue is present in all teleostean sequences except common carp Cyprinus carpio [5,8,[33][34][35]. Unlike its c-type counterpart, starry flounder g-type lysozyme has no potential cysteine residues capable of disulfide bridging, which is congruent with most other piscine g-type orthologues [4,5,7,8,[33][34][35][36].…”

“…Other teleostean fish, in common with starry flounder g-type, showed broad expression of the g-type in multiple tissues including hematopoietic and mucosal tissues exposed to external environment [8,[33][34][35]42]. For instance, in European seabass Dicentrarchus labrax L. and Asian seabass, the highest transcript levels of g-type lysozymes were found in gills and intestine, followed by head kidney [8,33], and in turbot S. maximus and Atlantic salmon Salmo salar, the highest levels were found in heart, spleen, and head kidney, respectively [34,42]. Hence, the basal tissue expression patterns of g-type lysozyme mRNAs might be species-specific.…”