Lysozymes in molluscs

Invertebrates lack antibody-mediated humoral immune systems; however, they are believed to possess efficient host defense mechanisms involving humoral defense molecules that are similar in function to antibodies. Lysozymes are a group of enzymes that cleave the glycosidic bonds between N-acetylmuramic acid and N-acetylglucosamine (two amino sugars) in the peptidoglycans that form bacterial cell walls. In bivalves, lysozymes are especially important antibacterial molecules because of their bactericidal ability. Recently, the presence of multiple lysozymes has been found in several species of bivalve molluscs such as the blue mussel, Mytilus edulis, and the eastern oyster, Crassostrea virginica. Therefore, to determine the molecular and biochemical properties of bivalve lysozymes, we have identified the cDNA sequences of three different lysozymes (CGL-1, -2, and -3) from the Pacific oyster, C. gigas, and have produced recombinant lysozymes (rCGL) using the methylotrophic yeast Pichia pastoris. The lysozyme CGL-1 mRNA was expressed in all tissues except for those of the adductor muscle. In contrast, CGL-2 gene was only expressed in digestive diverticula. Interestingly, in digestive diverticula, CGL-1 gene expression was detected in the same digestive cells as that of CGL-2. It is therefore possible that CGL-1 and CGL-2 play complementary roles in digestive organs. In situ hybridization revealed that CGL-3 mRNA was highly expressed in the mantle and haemocytes. These results suggest that CGL-1 and -2 serve as digestive enzymes for enteric and engulfed bacteria in digestive organs and that CGL-3 is involved in biodefense against invading microbes. Based on the results from experiments using recombinant lysozymes, we found significant differences among the characteristics of the three lysozymes, suggesting that these lysozymes have different functions in C. gigas.