title: Evidence for an angiotensin-(1-7) neuropeptidase expressed in the brain medulla and CSF of sheep.
abstract: Angiotensin-(1-7) [Ang-(1-7)] is an alternative product of the brain renin-angiotensin system that exhibits central actions to lower blood pressure and improve baroreflex sensitivity. We previously identified a peptidase that metabolizes Ang-(1-7) to the inactive metabolite product Ang-(1-4) in CSF of adult sheep. This study purified the peptidase 1445-fold from sheep brain medulla and characterized this activity. The peptidase was sensitive to the chelating agents o-phenanthroline and EDTA, as well as the mercury compound p-chloromercuribenzoic acid (PCMB). Selective inhibitors to angiotensin-converting enzyme, neprilysin, neurolysin, and thimet oligopeptidase did not attenuate activity; however, the metallopeptidase agent JMV-390 was a potent inhibitor of Ang-(1-7) hydrolysis (Ki = 0.8 nM). Kinetic studies using (125) I-labeled Ang-(1-7), Ang II, and Ang I revealed comparable apparent Km values (2.6, 2.8, and 4.3 μM, respectively), but a higher apparent Vmax for Ang-(1-7) (72 vs. 30 and 6 nmol/min/mg, respectively; p < 0.01). HPLC analysis of the activity confirmed the processing of unlabeled Ang-(1-7) to Ang-(1-4) by the peptidase, but revealed < 5% hydrolysis of Ang II or Ang I, and no hydrolysis of neurotensin, bradykinin or apelin-13. The unique characteristics of the purified neuropeptidase may portend a novel pathway to influence actions of Ang-(1-7) within the brain. Angiotensin-(1-7) actions are mediated by the AT7 /Mas receptor and include reduced blood pressure, decreased oxidative stress, enhanced baroreflex sensitivity, and increased nitric oxide (NO). Ang-(1-7) is directly formed from Ang I by neprilysin (NEP). We identify a new pathway for Ang-(1-7) metabolism in the brain distinct from angiotensin-converting enzyme-dependent hydrolysis. The Ang-(1-7) endopeptidase (A7-EP) degrades the peptide to Ang-(1-4) and may influence central Ang-(1-7) tone.
authors.affiliation: Hypertension and Vascular Research Center, Wake Forest School of Medicine, Winston Salem, North Carolina, USA.
authors.forename: Allyson C
authors.initials: AC
authors.lastname: Marshall
authors.forename: Nancy T
authors.initials: NT
authors.lastname: Pirro
authors.forename: James C
authors.initials: JC
authors.lastname: Rose
authors.forename: Debra I
authors.initials: DI
authors.lastname: Diz
authors.forename: Mark C
authors.initials: MC
authors.lastname: Chappell
mesh_terms: D000803:Angiotensin I / Q000096:biosynthesis* / Q000134:cerebrospinal fluid*; D000818:Animals; D001920:Bradykinin / Q000378:metabolism; D002847:Chromatography, Agarose; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D007700:Kinetics; D008526:Medulla Oblongata / Q000201:enzymology*; D017669:Mercury Compounds / Q000494:pharmacology; D009496:Neurotensin / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000134:cerebrospinal fluid*; D007703:Peptidyl-Dipeptidase A / Q000096:biosynthesis* / Q000134:cerebrospinal fluid*; D011480:Protease Inhibitors / Q000494:pharmacology; D012756:Sheep; D013379:Substrate Specificity
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D017669:Mercury Compounds; D009842:Oligopeptides; D010446:Peptide Fragments; D011480:Protease Inhibitors; C495199:apelin-13 peptide; C078372:JMV 390-1; D009496:Neurotensin; D000803:Angiotensin I; D007703:Peptidyl-Dipeptidase A; C118790:angiotensin I (1-7); D001920:Bradykinin
keywords: Ang-(1-7); Renin-angiotensin system; metabolism; neuropeptidase