Purification and partial characterization of a heat-resistant, cytosolic neuropeptidase from rat liver

A cholecystokinin octapeptide (CCK-8)-degrading peptidase was purified from rat liver cytosol by heat precipitation of other proteins followed by gel filtration, ion exchange chromatography and preparative gel electrophoresis, using a silicate binding assay to quantitate the degradation of radiolabeled CCK-8. The purified peptidase (M_r ˜ 60,000) had a pH optimum of 6.0; its activity was inhibited by EDTA and 1, 10-phenanthroline but not by phosphoramidon, calpain inhibitor I, bestatin or bacitracin. CCK-8 peptidase rapidly degraded radiolabeled Metenkephalin as well as ¹²⁵I-CCK-8, but not a series of other unrelated peptides. Unlabeled Leuenkephalin, β-casomorphin and neurotensin competitively inhibited the degradation of ¹²⁵I-CCK-8, suggesting that these opioids are also substrates for the enzyme. These data suggest that this protein is a novel hepatic enzyme which may play a role in the degradation of neuropeptides.