Hydrolysis of the C-terminal octapeptide of cholecystokinin by rat kidney membranes: characterization of the cleavage by solubilized endopeptidase-24.11.
Rat kidney membranes were solubilized by Triton X-100 and the CCK-8 degrading peptidases were resolved by chromatography on DEAE-cellulose. Four proteases were detected: two phosphoramidon-sensitive endopeptidases (EC 3.4.24.11), a bestatin-sensitive aminopeptidase and an unidentified enzyme. The pattern of cleavage of CCK-8 and shorter C-terminal fragments by endopeptidase 24.11 was investigated and indicated that the Gly29-Trp30, Trp30-Met31 and Asp32-Phe33 were scissile bonds. However, the cleavage pattern differed markedly from one CCK peptide to another: in the penta- and hexapeptide of CCK the bonds hydrolyzed were either Asp-Phe and Trp-Met or, Asp-Phe and Gly-Trp, respectively. The presence of the sulfate group on the tyrosine residue of CCK-8 influence markedly the nature of the major cleavage fragments produced by the endopeptidase. The major bonds cleaved were Asp-Phe, Trp-Met and Gly-Trp for unsulfated CCK-8, whilst for the sulfated octapeptide, the Trp-Met bond became a minor cleavage site.
D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D010450:Endopeptidases / Q000378:metabolism*; D006020:Glycopeptides / Q000494:pharmacology; D006868:Hydrolysis; D066298:In Vitro Techniques; D007668:Kidney / Q000201:enzymology*; D008566:Membranes / Q000201:enzymology; D015260:Neprilysin; D010446:Peptide Fragments / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats; D012844:Sincalide / Q000378:metabolism*; D012995:Solubility
D006020:Glycopeptides; D010446:Peptide Fragments; D011480:Protease Inhibitors; D010450:Endopeptidases; D015260:Neprilysin; D012844:Sincalide; C008890:phosphoramidon