Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.11 (CD10)
 9,792 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study was designed to evaluate the role of neutral endopeptidase (NEP) in modulating the airway smooth muscle contraction induced by endothelin-1 in isolated segments of guinea-pig trachea. Endothelin-1 (10(-9)-10(-6) M) produced a concentration-dependent contraction that reached a maximum by 30 min. The NEP inhibitor leucine-thiorphan (10(-5) M) significantly increased the contractile response to endothelin-1. The addition of leucine-thiorphan to tracheal segments precontracted by 10(-9) and 10(-8) M endothelin-1 increased isometric tension by 181 +/- 65% (mean +/- 1 S.E.M.; P less than 0.05) and by 138 +/- 49% (P less than 0.05), respectively. In contrast, the kininase II inhibitor captopril and the peptidase inhibitors leupeptin and bestatin had no effect. Preincubation of endothelin-1 with 1 microgram recombinant human NEP decreased the contractile activity of endothelin-1 by 72 +/- 9%, whereas no effect was observed using heat-inactivated NEP. We conclude that NEP modulates endothelin-induced contraction of airway smooth muscle in the guinea-pig trachea.
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PMID:Neutral endopeptidase modulates endothelin-1-induced airway smooth muscle contraction in guinea-pig trachea. 143 68

Endothelin-1 (ET-1) is a potent vasoconstrictor peptide secreted by endothelial cells. We investigated whether polymorphonuclear neutrophils (PMN) were able to destroy this peptide by enzymatic hydrolysis produced either by the membrane-bound endopeptidase 24.11 or by lysosomal proteinases released in the medium by activated cells. Resting and activated PMN were incubated with 125I-labelled ET-1 and the degradation fragments were analyzed by HPLC. A marked degradation of ET-1 was observed only in the presence of the stimulated cells, leading to the generation of seven radiolabelled peaks. Addition of phosphoramidon had no effect on the appearance of these metabolites, while soybean trypsin inhibitor abolished almost completely the degradation of the peptide, suggesting a role of cathepsin G in ET-1 hydrolysis. Among the purified leukocyte enzymes tested, cathepsin G hydrolyzed 125I-labelled ET-1 at the higher rate and gave rise to two radiolabelled peaks already observed in the presence of activated PMN. Incubation of unlabelled ET-1 with purified cathepsin G allowed to identify a major cleavage site corresponding to the His16-Leu17 bond, leading to the formation of inactive [1-16] fragments and the C-terminal pentapeptide. This mechanism of ET-1 inactivation could play a role in acute inflammatory reaction where PMN adhere to the vascular endothelial cells.
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PMID:Enzymatic degradation of endothelin-1 by activated human polymorphonuclear neutrophils. 147 5

The breakdown of endothelin-1 by crude membrane preparations of human kidney and choroid plexus was investigated. 125I-labeled endothelin-1 was degraded by both tissues in a phosphoramidon-sensitive way, suggesting a role of endopeptidase 24.11 in the in vitro metabolism of this peptide. Identification of the cleavage sites of purified human renal endopeptidase 24.11 in the sequence of endothelin-1 revealed that bonds involving the amino side of the hydrophobic amino acids (Ser4, Leu6, Val12, Phe14, His16, Leu17, Ile19) were susceptible to cleavage. Endothelin-1 appears thus to be degraded at multiple sites by endopeptidase 24.11 in vitro, producing inactive fragments.
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PMID:In vitro degradation of endothelin-1 by endopeptidase 24.11 (enkephalinase). 178 41

Endothelin-1 is a potent endothelium-derived vasoconstrictor and pressor peptide with uniquely sustained activity. We have examined the contribution of endogenously-generated endothelin-1 to the maintenance of basal vascular tone in healthy subjects. In these studies, on separate occasions, a combined inhibitor of endothelin converting enzyme (ECE) and neutral endopeptidase (NEP), phosphoramidon, a selective inhibitor of NEP, thiorphan, and a selective ETA receptor antagonist, BQ-123, were given via the brachial artery. Big endothelin-1, the precursor to endothelin-1, caused a slow onset dose-dependent forearm vasoconstriction, the magnitude of which was consistent with about 10% conversion to mature endothelin-1 in the forearm. Vasoconstriction to big endothelin-1 was abolished by co-infusion of phosphoramidon, whereas vasoconstriction to endothelin-1 was unaffected. Phosphoramidon caused progressive vasodilatation when infused alone, with blood flow increasing by 37% at 90 min (P = 0.02), whereas thiorphan caused vasoconstriction, consistent with NEP inhibition exerting its major effect on degradation of constrictor peptides, such as angiotensin and endothelin-1. Vasoconstriction to endothelin-1 was completely abolished by coinfusion of BQ-123, and BQ-123 alone produced progressive forearm vasodilatation, with blood flow increasing by 64% after 60 min (P = 0.001). These results demonstrate that endogenous production of endothelin-1 acts to sustain vascular tone in humans and indicate that ECE inhibitors and endothelium receptor antagonists may have therapeutic potential as vasodilators.
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PMID:Endogenous endothelin generation maintains vascular tone in humans. 747 28

Endothelin-1 (ET-1), a 21-residue vasoactive peptide, is produced in vascular endothelial cells from the 38-residue inactive intermediate big endothelin-1 via a specific cleavage at Trp-21-Val-22. The protease that catalyzes the conversion, endothelin-converting enzyme (ECE), constitutes a potential regulatory site for the production of the active peptide. We report the identification of ECE-1, a novel membrane-bound neutral metalloprotease that is expressed abundantly in endothelial cells in vivo and is structurally related to neutral endopeptidase 24.11 and Kell blood group protein. When transfected into cultured cells that normally secrete only big ET-1, the ECE-1 cDNA conferred the ability to secrete mature ET-1. In transfected cells, ECE-1 processes endogenously synthesized big ET-1 as well as exogenously supplied big ET-1, which interacts with ECE-1 on the cell surface. ECE-1 may provide a target for pharmacological intervention to alter ET-1 production.
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PMID:ECE-1: a membrane-bound metalloprotease that catalyzes the proteolytic activation of big endothelin-1. 806 89

1. In the anaesthetized, ganglion-blocked rat, intravenous boluses of endothelin-1, endothelin-2 and endothelin-3 induced a transient hypotensive effect followed by a potent long lasting pressor response (ED50 mmHg: 0.72 +/- 0.05, 1.8 +/- 0.2 and 2.7 +/- 0.3 nmol kg-1, respectively). The maximal effect for the three peptides was of a similar order of magnitude (delta MAP: 84 to 89 mmHg). Neither of these effects was influenced by phosphoramidon or thiorphan (10 mg kg-1, i.v.). 2. Intravenously administered big-endothelin-1 and -2 induced a transient (1-2 min) hypotension followed by a potent long lasting (> 25 min) vasopressor effect (ED50 mmHg: 1.8 +/- 0.2 and 6.7 +/- 0.4 nmol kg-1, respectively), with a similar maximal activity (delta MAP: 85 +/- 4 and 81 +/- 2.4 mmHg, respectively). The onset of the big-endothelin-1 vasopressor effect was more rapid (5-6 min) than that of big-endothelin-2 (10-13 min). Big-endothelin-3 was found to induce only a potent, long lasting (> 35 min) hypertension, with a maximal effect of 75 +/- 4.6 mmHg at 10 nmol kg-1 and an ED50 mmHg of 6.5 +/- 0.4 nmol kg-1. The onset of this effect was much slower (20-25 min) than that of the other proendothelins. Pressor responses induced by big-endothelin-1, -2 and -3 (3, 15 and 10 nmol kg-1, respectively) were markedly reduced (60, 80 and 100%) in the presence of phosphoramidon (10 mg kg-1, i.v.). Thiorphan (10 mg kg-1, i.v.) did not inhibit the effects of big-endothelin-1, -2 and -3. 3. In the electrically stimulated rat vas deferens, endothelin-I and -2 were found to be equipotent enhancers of the twitch response (EC100%: 4.0 +/- 0.4 nm and 7.9 +/- 4.8 nm, respectively), both about 3-4 fold as active as endothelin-3 (EC100%: 19 +/- 2.5 nM). Endothelin-1 and -3 showed a comparable maximalstimulatory effect (Emax: 296 +/- 30 and 262 +/- 24%) while endothelin-2 was less active (Emax: 194 +/- 30%).4. Big-endothelin-l and -2 were potent enhancers of the twitch response too (EC 100,%: 10.0 +/- 2.6 nM and 21.6 +/- 3.2 nM, respectively), with a comparable maximal stimulatory effect (Emax: 254 +/- 22 and 264 +/-24%). Big-endothelin-3 was found to be less potent (EC,100%: 275 +/- 21 nM), but retained a marked potentiating effect (Emax: 200 +/- 38%). Phosphoramidon, but not thiorphan, concentration-dependently(10 and 100 microM) reduced big-endothelin-1 (58 and 86% respectively) and big-endothelin-2 (21 and 56%)-mediated responses. Conversely, the big-endothelin-3 effect was reduced by phosphoramidon only at 100 microM (-70%), while thiorphan acts concentration-dependently (31 and 71% at 10 and 100 microM respectively); thus, in the rat vas deferens, big-endothelin-I and -2 were as potent as their corresponding endothelins, while big-endothelin-3 was about 20 times less potent than endothelin-3.5. The increasing effect of endothelin-2 (194 +/- 30% over baseline) was significantly enhanced by either 10 microM phosphoramidon (277 +/- 42%) or thiorphan (318 +/- 15%). The endothelin-I and endothelin-3-mediated twitch enhancement was not affected by the two protease inhibitors (10 microM).6. These results suggest that in vivo big-endothelin-1, -2 and -3, are processed through a similar phosphoramid on-sensitive enzymatic pathway although with different apparent affinity. This enzymatic process is probably attributable to a neutral endoprotease, distinct from neutral-endopeptidase 24.11(NEP). On the other hand, a NEP-like enzymatic activity may be involved, in the rat vas deferens, in the activation of big-endothelin-3 to endothelin-3 and in the metabolism of endothelin-2, but not of endothelin-I or endothelin-3.
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PMID:Comparison of the cardiovascular and neural activity of endothelin-1, -2, -3 and respective proendothelins: effects of phosphoramidon and thiorphan. 810 8

Endothelin-1 (ET-1), a 21 amino-acid potent vasoconstrictor peptide, is produced from the biologically inactive intermediate big ET-1 via an endoproteolytic cleavage between Trp-21 and Val-22 by endothelin converting enzyme (ECE). cDNA sequence analysis predicts that the two other members of the endothelin family, ET-2 and ET-3, are also generated from the corresponding intermediates called big ET-2 and big ET-3, respectively. The metalloproteinase inhibitor phosphoramidon inhibited the conversion of big ET-1 into mature ET-1 both in vivo and in cultured endothelial cells, suggesting that ECE may be a neutral metalloproteinase. In this study, we solubilized and partially purified ECE from the membrane fraction of porcine lung. Using gel filtration chromatography, we separated two distinct ECE activities, designated M1 (apparent molecular mass approx. 300 kDa) and M2 (approx. 65 kDa). Optimum pH for the cleavage of big ET-1 by M1 and M2 was 7.0 and 7.5, respectively. M1 efficiently converted human big ET-1(1-38) to ET-1, but not human big ET-2(1-37) or human big ET-3(1-41)-amide. In contrast, M2 converted both big ET-1 and big ET-2, but not big ET-3. M1 was inhibited by phosphoramidon (IC50 approx. 1 microM) but not by thiorphan or bacitracin. In contrast, M2 was inhibited by much lower concentrations of phosphoramidon (IC50 approx. 0.3 nM), as well as by thiorphan and bacitracin. ECE activity in M1 was able to bind to a concanavalin A-agarose column and was eluted by alpha-methyl-D-glucoside, indicating that the ECE is glycosylated. From these results, M1 and M2 from the porcine lung membrane are similar to the candidate of ECE in endothelial cells and neutral endopeptidase in kidney (EC 3.4.24.11), respectively. Taken in conjunction with the previous finding that neither thiorphan nor bacitracin affected the conversion of endogenously synthesized big ET-1 in cultured endothelial cells, we conclude that physiologically relevant ECE found in the endothelial cells is more similar to M1 than to M2.
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PMID:Characterization of phosphoramidon-sensitive metalloproteinases with endothelin-converting enzyme activity in porcine lung membrane. 843 80

Endothelin-1 (ET), the most potent vasoconstrictor yet discovered, is a peptide containing 21 amino acids with two intrachain disulfide bridges. With the aim of obtaining two-chain derivatives, Et was submitted to chemical and enzymatic treatments. Reaction of ET with CNBr in 70% HCOOH gave, in addition to the expected [Hse7 lactone]-7,8-seco-ET and unreacted material, a by-product whose molecular weight was 25 m.u. greater than that of ET. When the reaction mixture, after lyophilisation, was immediately quenched with NH3-saturated dry MeOH, two products could be recovered in a 5:1 ratio, both obtained by nucleophilic attack of the homoserine lactone: the expected [Hse7-NH2]-7,8-seco-ET and [Hse7]ET, resulting from competitive intramolecular reaction of the deprotonated alpha-amino group of the Asp8 residue. The Lys9-Glu10 bond turned out to be very resistant to enzymatic attack both by Lys-C-endopeptidase and trypsin. The 9,10-seco-ET derivative could be obtained by treatment with Lys-C-endopeptidase only by using a high enzyme/ET ratio and after a prolonged incubation time. Cleavage of the Lys9-Glu10 bond could not be achieved by treatment with trypsin, even with a high enzyme/substrate ratio. The main product was 13,14-seco-ET, deriving from the action of chymotripsin (present as an impurity in the trypsin preparation) on Tyr13. The structure of these peptides was confirmed by amino-acid sequence analysis and fast atom bombardment mass spectrometry (FAB-MS). Nicking of the ET structure at different positions had different impact on the biological properties of the resulting derivatives.
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PMID:Chemical and enzymatic treatment of endothelin. 856 76

1. We have studied the conversion of big endothelin-1 (big ET-1), big endothelin-2 (big ET-2) and big endothelin-3 (big ET-3) and characterized the enzyme involved in the conversion of the three peptides in guinea-pig lung parenchyma (GPLP). 2. Endothelin-1 (ET-1), endothelin-2 (ET-2) and endothelin-3 (ET-3) (10 nM to 100 nM) caused similar concentration-dependent contractions of strips of GPLP. 3. Big ET-1 and big ET-2 also elicited concentration-dependent contractions of GPLP strips. In contrast, big ET-3, up to a concentration of 100 nM, failed to induce a contraction of the GPLP. 4. Incubation of strips of GPLP with the dual endothelin converting enzyme (ECE) and neutral endopeptidase (NEP) inhibitor, phosphoramidon (10 microM), as well as two other NEP inhibitors thiorphan (10 microM) or SQ 28,603 (10 microM) decreased by 43% (P < 0.05), 42% (P < 0.05) and 40% (P < 0.05) the contractions induced by 30 nM of big ET-1 respectively. Captopril (10 microM), an angiotensin-converting enzyme inhibitor, had no effect on the contractions induced by big ET-1. 5. The incubation of strips of GPLP with phosphoramidon (10 microM), thiorphan (10 microM) or SQ 28,603 (10 microM) also decreased by 74% (P < 0.05), 34% and 50% (P < 0.05) the contractions induced by 30 nM big ET-2 respectively. As for the contractions induced by big ET-1, captopril (10 microM) had no effect on the concentration-dependent contractions induced by big ET-2. 6. Phosphoramidon (10 microM), thiorphan (10 microM) and SQ 28,603 (10 microM) significantly potentiated the contractions of strips of GPLP induced by both ET-1 (30 nM) and ET-3 (30 nM). However, the enzymatic inhibitors did not significantly affect the contractions induced by ET-2 (30 nM) in this tissue. 7. These results suggest that the effects of big ET-1 and big ET-2 result from the conversion to ET-1 and ET-2 by at least one enzyme sensitive to phosphoramidon, thiorphan and SQ 28,603. This enzyme corresponds possibly to EC 3.4.24.11 (NEP 24.11) and could also be responsible for the degradation of ETs in the GPLP.
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PMID:Role of the neutral endopeptidase 24.11 in the conversion of big endothelins in guinea-pig lung parenchyma. 882 61

Endothelin-1 (ET-1) by itself was not an effective stimulus for inducing superoxide (O2.) generation in human resting or DMSO-differentiated neutrophil-like HL-60 cells. ET-1 (0.01-100 nM) was not able to modulate O2. generation stimulated by the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP, EC50 = 4.24 +/- 1.63 nM in the absence and 3.16 +/- 1.95 nM in the presence of ET-1). Neither did ET-1 (0.01-100 nM) promote the mobilization of intracellular calcium ions or modulate fMLP-induced [Ca(2+)]i increase in this model of human neutrophils. Phosphoramidon, a neutral endopeptidase inhibitor, was not able to reveal any biological (O2.) or biochemical ([Ca(2+)]i response to ET-1 in the absence or in the presence of fMLP in these cells. These results indicate that DMSO-differentiated neutrophil-like HL-60 cells are not sensitive to ET-1 in terms of O2. generation or [Ca(2+)]i variations.
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PMID:Endothelin-1 does not modulate O2.release and [Ca(2+)]i variations in resting or differentiated HL-60 cells. 890 Apr 97


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