Vascular, Post Proline Cleaving Enzyme: Metabolism of Vasoactive Peptides
Vasoactive peptides contain a high proportion of proline residues which make them resistant to hydrolysis by many peptidases. However, post proline cleaving enzyme (PPCE; EC 126.96.36.199), a proline specific endopeptidase which specifically hydrolyzes internal peptide bonds on the carboxyl side of proline residues, has been shown to inactivate numerous vasoactive peptides including angiotensins, kinins, substance P, vasopressin and oxytocin. In order to determine whether PPCE could be involved in vascular metabolism of vasoactive peptides, we carried out localization and characterization studies of PPCE-like activity in hog aorta and mesenteric artery. PPCE was assayed fluorometrically at pH 7.0 using the specific PPCE substrate CBZ-Gly-Pro-4-methyl-coumarinylamide. The subcellular distribution of vascular PPCE was essentially the same as that of the cytosolic marker enzyme lactic dehydrogenase (LDH). PPCE was enriched six-fold in the cytosolic fraction (11.4 ± 2.7 units/mg) and, unlike the plasma membrane-bound proline specific exopeptidase dipeptidyl-(amino) peptidase IV (DAP IV; EC 188.8.131.52), little or no activity could be detected in the microsomal or plasma membrane fractions. Similar to PPCE characterized from other sites, vascular PPCE was stabilized and activated by dithiothreitol and EDTA, and inhibited by DFP, p-chloromercuriphenyl sulfonic acid, L-1-tosylamido-2- phenylethylchloromethyl ketone, Cu++, Ca++, and Zn++. Vascular PPCE was unaffected by inhibitors of trypsin and kallikrein (Aprotinin, ABTI), aminopeptidase M (bestatin, amastatin), neutral endopeptidase (phosphoramidon), angiotensin I converting enzyme (captopril) or carboxypeptidase N (MERGETPA). These data demonstrate that PPCE is present in vascular endothelium and/or smooth muscle. Although PPCE may participate in vascular metabolism of vasoactive peptides, its cytosolic localization and endopeptidase specificity suggests that its role is limited to peptide degradation after transport into the cell.
KeywordsProline Residue Plasma Membrane Fraction Neutral Endopeptidase Vasoactive Peptide Immunoelectrophoretic Analysis
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