Identification and characterization of the proteolytic enzymes in the developmental stages of the eel-pathogenic nematode Anguillicola crassus

Abstract

The proteolytic activities of homogenates prepared from the second larva (L2) and the third larva (L3) as well as the adult stage of the eel-pathogenic nematode Anguillicola crassus were examined using hemoglobin, azocoll, elastin-orcein, and keratin azure as substrates. Whole bodies of L2 larvae, the anterior third of the bodies of L3 larvae, and the anterior fifth of the bodies of adults were studied. Extracts of L2 contained a trypsin-like proteinase exhibiting a molecular weight of 38,000 Da on gelatin-substrate gel electrophoresis. The proteinase showed a pH optimum at 8 and activity against azocoll and keratin. An apparent molecular weight of 25,000 Da was determined for the trypsin-like proteinase of the L3. This enzyme possessed collagenolytic, keratinolytic and slight elastinolytic activity at an optimal pH of 8. Samples of adults contained an aspartyl proteinase with a molecular weight of 90,000 Da. When hemoglobin was used as the substrate, the enzyme displayed optimal activity at pH 5. It was concluded that the proteinases of the larval stages are penetration enzymes, whereas that of the adult stage is a digestive enzyme.