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)

Endothelins (ET) are a family of potent vasoactive peptides that are produced from biologically inactive intermediates, termed big endothelins, via a proteolytic processing at Trp21-Val/Ile22. We recently cloned and characterized a membrane-bound metalloprotease that catalyzes this proteolytic activation, endothelin-converting enzyme-1 (ECE-1) (Xu, D., Emoto, N., Giaid, A., Slaughter, C., Kaw, S., deWit, D., and Yanagisawa, M. (1994) Cell 78, 473-485). This enzyme was shown to function in the secretory pathway as well as on the cell surface. Here we report molecular cloning of another novel enzyme, ECE-2, that produces mature ET-1 from big ET-1 both in vitro and in transfected cells. The cDNA sequence predicts that bovine ECE-2 is a metalloprotease structurally related to ECE-1, neutral endopeptidase 24.11, and human Kell blood group protein. The deduced amino acid sequence of ECE-2 is most similar to ECE-1, with an overall identity of 59%. ECE-2 resembles ECE-1 in that it is inhibited in vitro by phosphoramidon and FR901533 but not by thiorphan or captopril, and it converts big ET-1 more efficiently than big ET-2 or big ET-3. However, ECE-2 also exhibits the following striking differences from ECE-1. (i) The sensitivity of ECE-2 to phosphoramidon is 250-fold higher as compared with ECE-1, while FR901533 inhibits both enzymes at similar concentrations. (ii) ECE-2 has an acidic pH optimum at pH 5.5, which is in sharp contrast to the neutral pH optimum of ECE-1. ECE-2 has a narrow pH profile and is virtually inactive at neutral pH. Chinese hamster ovary (CHO) cells, which lack detectable levels of endogenous ECE activity, secrete mature ET-1 into the medium when doubly transfected with ECE-2 and prepro-ET-1 cDNAs. However, ECE-2-transfected CHO cells do not efficiently produce mature ET-1 when present with an exogenous source of big ET-1 through coculture with prepro-ET-1-transfected CHO cells. These findings suggest that ECE-2 acts as an intracellular enzyme responsible for the conversion of endogenously synthesized big ET-1 at the trans-Golgi network, where the vesicular fluid is acidified.
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PMID:Endothelin-converting enzyme-2 is a membrane-bound, phosphoramidon-sensitive metalloprotease with acidic pH optimum. 779 12

Endothelin (ET) is a 21-residue potent vasoconstrictive peptide produced by vascular endothelial cells and formed from its precursor, big endothelin (big ET), by endothelin-converting enzyme (ECE). Here we report the cloning and functional expression of a complementary DNA encoding a rat ECE from endothelial cells. Rat ECE is a highly glycosylated protein consisting of 10 possible N-linked glycosylation sites, a zinc-binding domain, and a single membrane-spanning region. It has structural and sequence homology to neutral endopeptidase and Kell blood group protein, a putative neutral endopeptidase. Monoclonal antibody risen against purified rat lung ECE recognized broad glycosylated bands in membrane fractions prepared from both rat lung and COS cells transfected with a rat ECE expression vector. Expressed ECE was inhibited by phosphoramidon, but not by thiorphan. It also cleaved big ET-1 efficiently but not big ET-2 or big ET-3. Northern blot analysis revealed that ECE messenger RNA is widely expressed in many tissues.
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PMID:Cloning and functional expression of endothelin-converting enzyme from rat endothelial cells. 803 69

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

The two human endothelin-converting enzyme (ECE-1) isoforms, which differ by their N-terminal region, are encoded by a single gene. The gene is composed of 19 exons that span more than 68 kilobases and has been mapped to the 1p36 band of the human genome. The two isoform mRNAs display different tissue distributions. Their precursors are transcribed from two distinct start sites, upstream from exon 1 and exon 3, respectively. Sequence analysis of the two putative promoters revealed the presence of motifs characteristic for several transcription factors. Comparison of the ECE-1 gene structure with those of other zinc metalloproteases, as well as a phylogenetic study, confirm the existence of a metalloprotease subfamily composed of ECE-1, ECE-2, neutral endopeptidase, Kell blood group protein, and two bacterial enzymes.
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PMID:Organization of the gene encoding the human endothelin-converting enzyme (ECE-1). 853 Mar 72

Endothelin-converting enzyme-1 (ECE-1) is a type II integral membrane protein that belongs to a family of metalloproteases which includes ECE-2, neprilysin (neutral endopeptidase 24.11, EC 3.4.24. 11), and Kell blood group protein. ECE-1 cleaves its biologically inactive native substrate, big endothelin-1, to generate a powerful vasoactive 21-amino acid peptide, endothelin-1. ECE-1 consists of a short N-terminal cytoplasmic tail, a transmembrane hydrophobic domain, and a large extracellular domain containing the catalytic site with a conserved Zn-binding motif. We have constructed a secreted, soluble form of ECE-1 (solECE-1) by fusing the cleavable N-terminal signal sequence of human alkaline phosphatase in frame with the entire extracellular domain of ECE-1. Stable transfectant CHO cell lines expressing up to 6.1 mg of solECE-1 per liter culture medium were established and solECE-1 was purified to homogeneity using three chromatographic steps with a 24% yield. SolECE-1 behaves as a dimer of 110-kDa subunits. SolECE-1 has a sharp pH optimum, similar to the native form, ECE-1a, but has a slightly more acidic pH optimum of 6.1-6.4 than that of 6.7-6.9 for ECE-1a. At its optimal pH of 6.4, solECE-1 cleaved big ET-1:big ET-2:big ET-3 in a ratio of 8.1:1:1.4, was inhibited by phosphoramidon with an IC50 value of 0.35 +/- 0.05 microM, had a Km value of 4.65 +/- 0.78 microM for big ET-1, and had a kcat value of 5.82 +/- 0.21 min-1, all values comparable to those for ECE-1a at its optimal pH of 6.8. Phosphoramidon inhibition of both ECE-1a and solECE-1 is highly pH-dependent. At pH 5.8, phosphoramidon inhibited ECE-1a and solECE-1 with IC50 values of 14 and 33 nM, respectively, which are 49- and 1224-fold more potent than at pH 7.2. SolECE-1 is highly glycosylated, similar to ECE-1a. Deglycosylation of solECE-1 by peptide N-glycosidase F shifted the apparent molecular weight of solECE-1 to approximately 80 kDa and the deglycosylated form(s) of solECE-1 preserved at least 72% of the activity of the glycosylated form.
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PMID:Soluble human endothelin-converting enzyme-1: expression, purification, and demonstration of pronounced pH sensitivity. 980 68

Kell blood group protein shares a consensus sequence (H.E.X.X.H) with a large family of zinc-dependent endopeptidases. Kell has closest homology with neutral endopeptidase 24.11, endothelin converting enzyme-1 (ECE-1), and the PEX gene product that, as a group, comprise the M13 subfamily of mammalian neutral endopeptidases. The proteolytic activity of the M13 members, but not of Kell, has been previously demonstrated. A secreted form of wild-type Kell protein (s-Kell), devoid of the intracellular and transmembrane domains, was expressed in sf9 cells. As a negative control, an inactive mutant Kell protein (E582G) was expressed. As determined by N-terminal amino acid sequencing and mass spectrometry of the cleaved products, wild-type s-Kell, but not the control mutant protein, specifically cleaved big endothelin-3 (ET-3) at Trp(21)-Ile(22), yielding ET-3, and, to a much lesser extent, also cleaved big ET-1 and big ET-2 at Trp(21)-Val(22), yielding ET-1 and ET-2. Enzymatic activity was partially inhibited by phosphoramidon. s-Kell has an acidic pH optimum (pH 6.0 to 6.5). Like the recombinant protein, red blood cells of common Kell phenotype also preferentially process big ET-3, in contrast to Ko (null) cells that do not. These data demonstrate that the Kell blood group protein is a proteolytic enzyme that processes big ET-3, generating ET-3, a potent bioactive peptide with multiple biological roles.
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PMID:Proteolytic processing of big endothelin-3 by the kell blood group protein. 1043 32

Because of their roles in controlling the activity of several bio-active peptides, members of the neprilysin family of zinc metallopeptidases have been identified as putative targets for the design of therapeutic agents. Presently, six members have been reported, these are: neprilysin, endothelin-converting enzyme (ECE)-1 and ECE-2, the Kell blood group protein, PHEX (product of the phosphate-regulating gene with homologies to endopeptidase on the X chromosome) and X-converting enzyme (XCE). In order to identify new members of this important family of peptidases, we designed a reverse transcriptase-PCR strategy based on conserved amino acid sequences of neprilysin, ECE-1 and PHEX. We now report the cloning from mouse testis of a novel neprilysin-like peptidase that we called NL1. NL1 is a glycoprotein that, among the members of the family, shows the strongest sequence identity with neprilysin. However, in contrast with neprilysin and other members of the family which are type II integral membrane proteins, NL1 was secreted when expressed in cultured mammalian cells, likely due to cleavage by a subtilisin-like convertase at a furin-like site located 22 amino acid residues in the C-terminus of the transmembrane domain. The recombinant enzyme exhibited neprilysin-like peptidase activity and was efficiently inhibited by phosphoramidon and thiorphan, two inhibitors of neprilysin. Northern blot analysis and in situ hybridization showed that NL1 mRNA was found predominantly in testis, specifically in round and elongated spermatids. This distribution of NL1 mRNA suggests that it could be involved in sperm formation or other processes related to fertility.
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PMID:Molecular cloning and biochemical characterization of a new mouse testis soluble-zinc-metallopeptidase of the neprilysin family. 1074 71

The Kell blood group protein is a zinc endopeptidase that yields endothelin-3, a potent bioactive peptide, by cleavage of big endothelin-3, a larger intermediate precursor. On red cells, Kell protein is linked by a single disulfide bond to XK, a protein that traverses the membrane 10 times and whose absence, as occurs in the McLeod phenotype, is associated with a set of clinical symptoms that include nerve and muscle disorders and red cell acanthocytosis. Previous studies indicated that Kell is primarily expressed in erythroid tissues, whereas XK has a wider tissue distribution. The tissue distribution of Kell protein has been further investigated by Northern blot analysis, PCR-screening of tissue complementary DNAs (cDNAs), and Western immunoblots. Screening of an RNA dot-blot panel confirmed that Kell is primarily expressed in erythroid tissues but is also expressed in a near equal amount in testis, with weaker expression in a large number of other tissues. PCR-screening of cDNAs from different tissues and DNA sequencing of the products gave similar results. In 2 of the nonerythroid tissues tested, testis and skeletal muscle, Kell protein was detected by Western immunoblotting. In skeletal muscle, isolation of XK with a specific antibody coisolated Kell protein. These studies demonstrate that Kell is expressed in both erythroid and nonerythroid tissues and is associated with XK.
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PMID:Expression of Kell blood group protein in nonerythroid tissues. 1089 71