Isolation and characterization of the digestive proteinases in the earthworm Lumbricus terrestris Linnaeus

Abstract

1. 1. Digestive organ homogenates and intestinal fluid from Lumbricus demonstrated strong activity on the synthetic substrates TAME and BAPA; a pH optimum at 8.35–8.45 was found for both substrates. The TAME components were found to be stable at pH 4.5–8.0. 2. 2. The TAME and BAPA activities were inhibited by ovumucoid and DFP, suggesting the presence of a tryptic-like serine proteinase. 3. 3. The crude extracts demonstrated strong carboxypeptidase-A activity on the synthetic substrate HPLA, with pH optima at 5.2 and 7.7. The carboxypeptidase-A activity was inhibited 30 per cent with EDTA and 1,10-phenanthroline. Crude extracts demonstrated activity on Bz-GL, with a pH optimum at 7.7–8.0. 4. 4. No enzymatic activity occurred with the synthetic substrates l-leucinamide and GPANA. 5. 5. Thirteen protein components were separated by acrylamide gel electrophoresis of which five demonstrated caseinase activity. 6. 6. The first and second isolated proteolytic components did not hydrolyze any of the synthetic substrates tested. 7. 7. Proteolytic components 3 and 5 hydrolyzed HPLA. Component 3 demonstrated pH optima of 5.2 and 7.7, while component 5 had a pH optimum of 7.7 only. 8. 8. Proteolytic component 4 hydrolyzed TAME and BAPA. Component 4 also hydrolyzed Bz-GL, the amino side of the basic amino acid. 9. 9. A sixth component hydrolyzed Bz-GL, but caseinase activity was not apparent.