Hydrolysis of α-human atrial natriuretic peptide in vitro by human kidney membranes and purified endopeptidase-24.11. Evidence for a novel cleavage site

Abstract

alpha-Human atrial natriuretic peptide (hANP) is secreted by the heart and acts on the kidney to promote a strong diuresis and natriuresis. In vivo it has been shown to be catabolized partly by the kidney. Crude microvillar membranes of human kidney degrade 125I-ANP at several internal bonds generating metabolites among which the C-terminal fragments were identified. Formation of the C-terminal tripeptide was blocked by phosphoramidon, indicating the involvement of endopeptidase-24.11 in this cleavage. Subsequent cleavages by aminopeptidase(s) yielded the C-terminal dipeptide and free tyrosine. Using purified endopeptidase 24.11, we identified seven sites of hydrolysis in unlabelled alpha-hANP: the bonds Arg-4-Ser-5, Cys-7-Phe-8, Arg-11-Met-12, Arg-14-Ile-15, Gly-16-Ala-17, Gly-20-Leu-21 and Ser-25-Phe-26. However, the bonds Gly-16-Ala-17 and Arg-4-Ser-5 did not fulfil the known specificity requirements of the enzyme. Cleavage at the Gly-16-Ala-17 bond was previously observed by Stephenson & Kenny [(1987) Biochem. J. 243, 183-187], but this is the first report of an Arg-Ser bond cleavage by this enzyme. Initial attack of alpha-hANP by endopeptidase-24.11 took place at a bond within the disulphide-linked loop and produced a peptide having the same amino acid composition as intact ANP. The bond cleaved in this metabolite was determined as the Cys-7-Phe-8 bond. Determination of all the bonds cleaved in alpha-hANP by endopeptidase-24.11 should prove useful for the design of more stable analogues, which could have therapeutic uses in hypertension.

Natriuretic Peptide System Gene Expression in Human Coronary Arteries

“…It can be anticipated that because NPs antagonize the action of the renin-angiotensin system (RAS) (de Bold and Bruneau 2000), circulating or locally expressed NPs would modulate the cardiovascular growth processes that are dependent on locally expressed or circulating systems, such as the RAS, and that are present in the vasculature (Linz et al 1989). For example, ANF is capable of inhibiting thymidine incorporation by rat mesangial cells (Appel 1988;Vanneste et al 1988), and vasodilators that are capable of elevating intracellular cGMP levels, such as sodium nitroprusside and ANF, inhibit the proliferation of cultured SMCs (Kariya et al 1989). In addition, ANF can inhibit, in a dose-dependent manner, PDGFstimulated thymidine incorporation by SMCs (Abell et al 1989) and may act as an anti-growth factor for endothelial cells .…”

Hydrolysis of iodine labelled urodilatin and ANP by recombinant neutral endopeptidase EC. 3.4.24.11

“…Several studies demonstrated that NEP is the main enzyme that degrades ANP, through a single cleavage between cysteine-105 and phenylalanine-106 (Olins et al, 1987;Stephenson & Kenny, 1987). The product of the initial proteolytic attack is subject to further cleavage within the loop, and at the amino-and carboxy-termini (Olins et al, 1986;Stephenson & Kenny, 1987;Vanneste et al, 1988). The different stability of ANP and URO to degradation may account for the differences in the intensity of their renal effects.…”

Angiotensin receptor/neprilysin inhibitor: a novel therapy in the treatment of heart failure

“…[12][13][14] Neprilysin has been called by many other names, including enkephalinase, neutral endopeptidase, vasopeptidase and atriopeptidase. [10][11][12][13][14][15] Degradation by neprilysin is one of the two major means of elimination of natriuretic peptides, the other being through a clearance receptor (the natriuretic peptide clearance receptor; NPRC or NPR3). 16 Inhibition of neprilysin, therefore, represents a major approach in enhancing endogenous natriuretic peptide levels and activity.…”

“…10,11 It hydrolyses atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (and possibly urodilatin). [12][13][14] Neprilysin has been called by many other names, including enkephalinase, neutral endopeptidase, vasopeptidase and atriopeptidase.…”