Molecular isolation and characterization of a haemocyanin ofMacrobrachium rosenbergiireveal its antibacterial activities

Abstract

Haemocyanin is a multi-subunit protein complex found in the haemolymph and is involved in the immune system of crustaceans. In this study, a haemocyanin gene of Macrobrachium rosenbergii, designated MrHc, was successfully isolated. The MrHc gene contained an open reading frame (ORF) of 1,992 nucleotides, encoding a protein of 663 amino acid residues with a molecular mass of 76.5 kDa. The deduced amino acid sequence contained distinct structural motifs of the haemocyanin superfamily, including an all-alpha domain, a copper-containing domain and an immunoglobulin-like domain. Based on the phylogenetic analysis, the MrHC protein demonstrated a close relationship with the haemocyanins of Palaemon carinicauda and Macrobrachium nipponense. The MrHc gene was expressed in various shrimp tissues, including the hepatopancreas, gill, haemocytes, stomach and muscle. After Macrobrachium rosenbergii nodavirus (MrNV) challenge tests, the MrHc gene was up-regulated 237-fold at day 2. A recombinant protein of the MrHc immunoglobulin-like domain exhibited antibacterial activity against Vibrio vulnificus, V. parahaemolyticus, Aeromonas caviae, A. veronii, A. hydrophila and Bacillus cereus. This study suggested that MrHc may play important roles in the shrimp innate immune response to MrNV infection and bacterial infection.