Angiotensin processing is partially carried out by carboxypeptidases in the rat mesenteric arterial bed perfusate.
Here we investigated the possible association between the carboxypeptidase A (CPA)-like activity of the rat mesenteric arterial bed (MAB) perfusate and the ability of this fluid of forming angiotensin (Ang) 1-9 and Ang 1-7 upon incubation with Ang I and Ang II, respectively. Initially, we observed that anion exchange chromatography of the perfusate would consistently split the characteristic Z-Val-Phe-hydrolyzing activity of CPA-like enzymes into five distinct peaks, whose proteolytic activities were then determined using also Ang I and Ang II as substrates. The resulting proteolytic profile for each peak indicated that rat MAB perfusate contains a complex mixture of carboxypeptidases; tentatively, five carboxypeptidases were distinguished based on their substrate preferences toward Z-Val-Phe, Ang I and Ang II. The respective reactions, namely, Z-Val-Phe cleavage, Ang I to Ang 1-9 conversion and Ang II to Ang 1-7 conversion, were inhibited by 1,10-phenanthroline and nearly fully blocked by potato carboxypeptidase inhibitor. Also, all the CPA-like activity peaks prepared by anion exchange chromatography were tested negative for contaminating Ang I-converting enzyme-2, cathepsin A and prolylcarboxypeptidase. Overall, our results indicate that rat MAB perfusate contains a multiplicity of Ang I and Ang II-processing CPA-like enzymes whose proteolytic specificities suggest they might perform peculiar regulatory roles in the local renin-angiotensin system.
Departamentos de Bioquímica e Imunologia, da Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Brazil.
Hugo J V
HJ
Pereira
Laura L
LL
Souza
Maria Cristina O
MC
Salgado
Eduardo B
EB
Oliveira
D000803:Angiotensin I / Q000378:metabolism; D000804:Angiotensin II / Q000378:metabolism; D000809:Angiotensins / Q000378:metabolism*; D000818:Animals; D002268:Carboxypeptidases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008638:Mesenteric Arteries / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010477:Perfusion; D010618:Phenanthrolines / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D051381:Rats; D017208:Rats, Wistar; D013379:Substrate Specificity
D000809:Angiotensins; D010446:Peptide Fragments; D010618:Phenanthrolines; D011480:Protease Inhibitors; C506331:angiotensin I (1-9); D000804:Angiotensin II; D000803:Angiotensin I; D002268:Carboxypeptidases; C118790:angiotensin I (1-7); C025205:1,10-phenanthroline