Localization of chitinolytic enzymes in blood of turbot, Scophthalmus maximus, and their possible roles in defence

The intracellular localizations ofchitinase and β-N-acetylglucosaminidase were detected in turbot blood smears, using a novel method employing fluorogenic substrates. The two enzymes showed different distributions, with chitinase being more generally distributed and N-acetylglucosaminidase being strongly associated with distinct intracellular bodies, probably lysosomes. The fluorogenic substrates were used to analyse soluble and membrane fractions of homogenates of red and white blood cells prepared on Percoll gradients. In the leucocytes, the chitinase and N-acetylglucosaminidase activities were mostly in the soluble fraction. In the erythrocytes the activities were lower, at about one-hundredth and one-tenth specific activities, respectively, and were distributed between soluble and membrane-bound fractions at about 2 : 1 and 3 : 1, respectively. In contrast, lysozyme had a soluble distribution in leucocytes and was not detected in erythrocytes. Plasma was rich in chitinase and lysozyme activities but had no detectable N-acetylglucosaminidase. Two possible roles for the chitinolytic enzymes are discussed: defence against pathogens and processing of glycoproteins or glucosaminoglycans. Evidence for a defence role for the chitinase and lysozyme is provided by demonstrating that they had inhibitory activity against the chitinous fungus Mucor mucedo .