Identification and phylogenetic analysis on lipopolysaccharide and β-1,3-glucan binding protein (LGBP) of kuruma shrimp Marsupenaeus japonicus

Summary: A lipopolysaccharide (LPS) and β-1,3-glucan binding protein (LGBP) gene was cloned from hemocytes of kuruma shrimp Marsupenaeus japonicus by reverse-transcription polymerase chain reaction (RT-PCR), cloning and sequencing of overlapping PCR, and rapid amplification of cDNA ends (RACE) method. The open reading frame (ORF) of M. japonicus LGBP is 1062bp and encodes a 354 amino acid (aa) sequence with a 23aa signal peptide. The calculated molecular mass of the mature protein (331aa) is 40.15kDa with an estimated pI of 4.78. The M. japonicus LGBP sequence contains (1) two putative N-linked glycosylation sites, (2) two putative integrin-binding motifs, (3) a kinase C phosphorylation site (KCPS), (4) a glucanase motif (GM), and (5) two potential polysaccharide recognition motifs (polysaccharide binding motif (PsBM) and β-glucan recognition motif (GRM)), and with features of tryptophan-rich, slight homology to lysozyme, and slight homology to lectin. A sequence comparison showed that the deduced amino acids of M. japonicus LGBP has an overall high similarity to penaeid LGBP and βGBP (85.6–89.9%), lobster Homarus gammarus βGBP (77.0%), and crayfish Pacifastacius leniusculus LGBP (67.8%). The phylogenetic analysis revealed that M. japonicus LGBP grouped together with other crustacean LGBP and βGBP, and was close to termite GNBP, but was far way from moth βGBP, βGRP, fly GNBP, and mosquito βGRP. The LGBP of M. japonicus was strongly expressed in hemocytes. The LGBP mRNA transcript in hemocytes of M. japonicus was significantly upregulated 12–48h after a LPS injection, indicating activation of the innate immune system through the binding of the LGBP and LPS complex. [Copyright &y& Elsevier]