Abstract:In this study, peptide PGRP (designated HdPGRP) was identified and characterized from the abalone Haliotis discus hannai. Multiple alignments and phylogenetic analyses strongly suggested that HdPGRP is a new member of the PGRP superfamily and belongs to the short PGRP family, similar to peptides from other marine mollusks. The full length of HdPGRP is 1467 bp, encoding a polypeptide of 354 amino acids (aa) with a signal peptide (1~18 aa), an SH3b domain (93~160 aa), a typical PGRP domain (179~322 aa), and an Ami_2 domain (191~322 aa). In addition, four conserved Zn2+-binding sites (H209, Y255, H318, and C330) and five conserved amide-catalysis sites (H209, Y255, H318, T328, and C330) were found in the HdPGRP sequence. In abalone, hdpgrp exhibited different tissue expression patterns, and was strongly expressed in the hepatopancreas, moderately expressed in hemocytes, mantle, and gills, and slightly expressed in muscle. Vibrio anguillarum is one of the main pathogens of H. discus hannai; after V. anguillarum infection, expression of hdpgrp in hemocytes showed a trend of first increasing and then decreasing, reaching a maximum at 24 h. Subsequently, expression of HdPGRP decreased, and there was no significant difference compared with the control group at 72 h, demonstrating that expression of HdPGRP had returned to normal levels. SDS-PAGE results showed that recombinant HdPGRP (rHdPGRP) has a molecular mass of 30 kDa, which is in line with the value predicted for HdPGRP. PGRPs usually have amidase activity, degrading peptidoglycan by hydrolyzing the amide bond that links peptide units to muramic acid residues of glycan strands. rHdPGRP exhibited Zn2+-dependent amidase activity and catalyzed the degradation of insoluble peptidoglycan. In addition, rHdPGRP exhibited significant antibacterial activity against the gram-positive bacterium Micrococcus luteus in the logarithmic phase in the presence of Zn2+, indicating that the antibacterial activity of HdPGRP might be dependent on amidase activity. In summary, HdPGRP plays an important role in PGRP-mediated antibacterial mechanisms, especially for eliminating invading bacteria.
