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)

The sequence of the amino-terminal portion of human parathyroid hormone, particularly the identity of residues 22, 28, and 30 (the subject of discrepancies in recent published reports), has been reexamined by two basic methods of structural analysis. A fresh lot of human parathyroid hormone isolated from pooled adenoma tissue was analyzed by Edman degradation with identification of critical residues by thin-layer chromatography and gas-liquid chromatography. In the second approach, -14C or tritiated amino acids were incorporated during biosynthesis of the human hormone in slices of parathyroid glands in vitro; the appropriate amino acid residues were then determined as the -14C or tritiated phenythiohydantoin derivatives of the amino acid after Edman degradation, or by peptide isolation after appropriate cleavage with endopeptidase, or both. The results confirm our previous findings that residue 22 is glutamic acid, residue 28 is leucine, and residue 30 is aspartic acid.
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PMID:A reinvestigation of the amino-terminal sequence of human parathyroid hormone. 112 1

The dynamics of parathyroid hormone (PTH) biosynthesis, storage, and secretion in bovine parathyroid slices in vitro in response to alterations in the concentrations of extracellular calcium were studied. Hormone biosynthesis was evaluated by using polyacrylamide gel electrophoresis to measure incorporation of [3H]leucine into newly synthesized PTH and proparathyroid hormone (ProPTH) during short (35 min) incubations. Amounts of newly synthesized PTH stored in and secreted from the tissue slices were determined by electrophoretic analysis of [3H]PTH in extracts of tissue and media. Total PTH and ProPTH is slices and media were measured by specific radioimmunoassays. PTH secretion rates changes 5-fold when calcium was lowered from 2mM to 1mM. Secretion of some PTH continued despite high concentrations of calcium (5 mM). Biosynthesis of ProPTH was changed only slightly, and conversion of ProPTH to PTH was independent of the extracellular calcium concentration. Tissue stores of PTH increased during incubation of parathyroid slices in medium containing high amounts of calcium. The increase in stores was much less, however, than predicted by the findings of marked suppression of secretion and little change in rates of PTH biosynthesis. In high concentrations of calcium, a large fraction (up to 50%) of newly synthesized PTH was degraded within the tissue, whereas in low concentrations of calcium, little (less than 10%) of the PTH was degraded. No fragments of PTH or ProPTH were identified in either extracts of tissue or media, suggesting that degradation occurred rapidly by general proteolysis rather than by limited, specific endopeptidase activity. The data suggest that the parathyroid cell contains a calcium-sensitive degradative pathway for PTH and that this pathway may be involved in the regulation of hormone production and secretion.
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PMID:Calcium-dependent intracellular degradation of parathyroid hormone: a possible mechanism for the regulation of hormone stores. 115 61

A membrane-bound metallo-endopeptidase that hydrolyzes human parathyroid hormone (1-84) and reduced hen egg lysozyme between hydrophilic amino acid residues was isolated from rat kidney [Yamaguchi et al. (1991) Eur. J. Biochem. 200, 563-571]. In this study, the hydrolyses of various peptide hormones and neuropeptides by the metallo-endopeptidase were examined using an automated gas-phase protein sequencer. The purified enzyme hydrolyzed the oxidized insulin B chain and substance P most rapidly, followed by big endothelin 1, neurotensin, angiotensin 1, endothelin 1, rat alpha-atrial natriuretic peptide and bradykinin, in this order. The enzyme mainly cleaved these peptides at bonds involving a hydrophilic amino acid residue. However, it cleaved bonds between less hydrophilic amino acid pairs in several short peptides, e.g. at the His5-Leu6 bond in oxidized insulin B chain, the Ile28-Val29 bond in big endothelin-1 and the Ile5-His6 and Phe8-His9 bonds in angiotensin 1. The enzyme cleavage sites of oxidized insulin B chain and angiotensin 1 were different from the reported sites cleaved by meprin and by endopeptidase 2, respectively. Kinetic determination of bradykinin hydrolysis by the purified enzyme yielded values of Km = 18.1 microM and kcat = 0.473 s-1, giving a ratio of kcat/Km = 2.62 x 10(4) s-1.M-1. The Km value was about 20-fold lower than that reported for meprin and endopeptidase 2. These results indicate that the membrane-bound metallo-endopeptidase from rat kidney is distinguished from meprin and endopeptidase 2 in its substrate specificity and is not parathyroid hormone specific, but has potential capacities to inactivate various biologically active peptide hormones and neuropeptides in vivo.
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PMID:A membrane-bound metallo-endopeptidase from rat kidney. Characteristics of its hydrolysis of peptide hormones and neuropeptides. 137 51

A glutamic acid-specific protease has been purified to homogeneity from Bacillus licheniformis ATCC 14580 utilizing Phe-Leu-D-Glu-OMe-Sepharose affinity chromatography and crystallized. The molecular weight of the protease was estimated to be approximately 25,000 by SDS-polyacrylamide gel electrophoresis. This protease, which we propose to call BLase (glutamic acid-specific protease from B. licheniformis ATCC 14580), was characterized enzymatically. Using human parathyroid hormone (13-34) and p-nitroanilides of peptidyl glutamic acid and aspartic acid, we found a marked difference between BLase and V8 protease, EC 3.4.21.9, although both proteases showed higher reactivity for glutamyl bonds than for aspartyl bonds. Diisopropyl fluorophosphate and benzyloxycarbonyl Leu-Glu chloromethyl ketone completely inhibited BLase, whereas EDTA reversibly inactivated the enzyme. The findings clearly indicate that BLase can be classified as a serine protease. To elucidate the complete primary structure and precursor of BLase, its gene was cloned from the genomic DNA of B. licheniformis ATCC 14580, and the nucleotide sequence was determined. Taking the amino-terminal amino acid sequence of the purified BLase into consideration, the clones encode a mature peptide of 222 amino acids, which follows a prepropeptide of 94 residues. The recombinant BLase was expressed in Bacillus subtilis and purified to homogeneity. Its key physical and chemical characteristics were the same as those of the wild-type enzyme. BLase was confirmed to be a protease specific for glutamic acid, and the primary structure deduced from the cDNA sequence was found to be identical with that of a glutamic acid-specific endopeptidase isolated from Alcalase (Svendsen, I., and Breddam, K. (1992) Eur. J. Biochem. 204, 165-171), being different from V8 protease and the Glu-specific protease of Streptomyces griseus which consist of 268 and 188 amino acids, respectively.
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PMID:Purification, characterization, cloning, and expression of a glutamic acid-specific protease from Bacillus licheniformis ATCC 14580. 142 18

A metallo-endopeptidase, which appears to be an integral membrane protein of rat kidney, was purified to homogeneity by a series of standard chromatographic procedures. This enzyme significantly hydrolyzed human parathyroid hormone [hPTH(1-84)] and a synthetic substrate Suc-Leu-Leu-Val-Tyr-Mec (Suc = succinyl, Mec = 4-methyl-coumarinyl-7-amide). The purified enzyme had apparent molecular masses of 250 kDa on gel filtration, and 88 kDa and 245 kDa on sodium dodecyl sulfate/polyacrylamide gel electrophoresis under reducing and non-reducing conditions, respectively. Its pH optimum for activity was 8.0-8.5 and its isoelectric point was pH 4.9. Its activity was inhibited by EDTA, EGTA and o-phenanthroline, but not by phosphoramidon. The metal-depleted enzyme was reactivated by the addition of metal ions. The enzyme was also inhibited by chymostatin and eglin C, and by thiol compounds. Of the synthetic substrates examined, the enzyme hydrolyzed only Suc-Leu-Leu-Val-Tyr-Mec, one of the synthetic substrates for alpha-chymotrypsin. It did not hydrolyze synthetic substrates with less than four amino acid residues with tyrosine in the P1 position. The enzyme hydrolyzed hPTH and reduced hen egg lysozyme but did not hydrolyze azocasein or [3H]methyl-casein. NH2-terminal amino acid sequence analyses of the degradation products of hPTH(1-84) and reduced hen egg lysozyme by the purified enzyme revealed that the enzyme preferentially cleaved these peptides at peptide bonds flanked by hydrophilic amino acid residues. Amino acid analyses showed that the main degradation products of PTH were hPTH(17-29), hPTH(30-38) and hPTH(74-84). The ability of the enzyme to hydrolyze peptide bonds flanked by hydrophilic amino acid residues and its inability to degrade azocasein distinguish it from several other kidney endopeptidases reported, such as endopeptidase 24.11 and meprin.
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PMID:A membrane-bound metallo-endopeptidase from rat kidney hydrolyzing parathyroid hormone. Purification and characterization. 188 19

An endopeptidase catalyzing cleavage of parathyroid hormone to specific C-terminal and N-terminal fragments was identified in a partially purified membrane fraction from rat kidney. Fractionation on sucrose gradient showed that this activity is present primarily in a light membrane fraction rather than in the basal-lateral membranes, or in the classic lysosomal fraction. The endopeptidase can be extracted from the membranes by freezing and thawing, it has an acid pH optimum, and it catalyzes production of specific fragments of PTH. The major C-terminal fragment produced has its N-terminus at residue 39 of the native hormone. This fragment is identical to the primary PTH fragment found in kidney tissue following injection of iodinated PTH into the systemic circulation of rats (D'Amour et al., 1979). Finally, the cleavage of PTH by this acid endopeptidase is activated by physiological concentration of ATP (10(-4) - 10(-3)M). These results suggest that this enzyme may be involved in PTH catabolism by the kidney, that it may be located in a specialized cell fraction and that hormone catabolism may be regulated by the energy status of the cell.
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PMID:Identification of an ATP-activated endopeptidase from rat kidney which catalyzes cleavage of parathyroid hormone to fragments identical to those produced in the rat kidney in vivo. 633 13

Data from several laboratories indicate that hepatic mechanisms may have a distinctive role in the metabolism of intact hormone after secretion, a process that accounts, at least partly, for the heterogeneity of circulating parathyroid hormone. Accordingly, we studied the proteolysis of intact hormone by isolated rat Kupffer cells and hepatocytes. Kupffer cells (10(6) cells/ml) and hepatocytes (10(7) cells/ml) were incubated with unlabeled and (125)I-labeled bovine parathyroid hormone at 37 degrees C for periods ranging up to 2 h. When incubated with Kupffer cells, intact hormone disappeared with a t((1/2)) of 12+/-4 min. Radio-immunoassays using sequence-specific antisera showed that the dominant hormonal fragments recovered in the medium have an apparent molecular weight of approximately 6,000, lack amino-terminal antigenic determinants, and react in assays that specifically recognize determinants in the carboxy-terminal portion of the intact hormone. Amino-terminal fragments also were detected in high concentrations, particularly after short incubation periods. Radioiodinated fragments resulting from incubation of (125)I-labeled bovine parathyroid hormone with Kupffer cells had the same apparent size as fragments derived from the metabolism of unlabeled, intact hormone; when analyzed by Edman degradation, positions 34 and 37 of the intact hormone sequence were the amino-terminal amino acids of these dominant carboxy-terminal fragments. Hepatocytes did not hydrolyze the hormone. Thus, metabolism of parathyroid hormone by Kupffer cells results in the appearance of fragments in the media that are immunochemically indistinguishable from, and chemically identical with, those found in plasma when intact hormone is injected intravenously. This indicates that the proteolysis observed in vitro accurately reflects the metabolism of the hormone in vivo. The detection of amino-terminal fragments in concentrations nearly equal to those of carboxy-terminal fragments indicates that cleavage of intact hormone is, initially, by an endopeptidase(s). Kupffer cells may be a source from which specific protease(s) that hydrolyze parathyroid hormone can be characterized, particularly in terms of enzymic specificity and requirements for inhibition. Detailed analysis of the cellular and molecular events during incubation of parathyroid hormone with these cells may help to clarify the biologic significance of the peripheral metabolism of the hormone.
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PMID:Metabolism of parathyroid hormone by isolated rat Kupffer cells and hepatocytes. 700 37

Cleavage of parathyroid hormone (PTH) is catalyzed by an endopeptidase associated with a partially purified membrane preparation from bovine kidney cortex. This enzyme was found to have an acid pH optimum and to be easily extracted from the membranes by a single freeze-thaw cycle. The cleavage is remarkable in that it appears to be restricted to a small region of the PTH peptide chain, generating fragments which are not further degraded. The dominant products are a large fragment, COOH-terminal in origin, and a small fragment from the NH2 terminus. The small fragment is biologically active and its activity establishes that it contains at least the first 29 amino acids in PTH. The large fragment has no biological activity. The cleavage of PTH was demonstrated both with iodinated PTH and with unlabeled hormone by immunoassay and by labeling the large fragment after its production. Microsequencing of the large fragment showed that, in fact, two products are produced: one with its NH2 terminus at position 38 of PTH and one with its NH2 terminus at position 35. These fragments are remarkably similar to those generated in both the liver and the kidney in vivo.
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PMID:Specific cleavage of bovine parathyroid hormone catalyzed by an endopeptidase from bovine kidney. 702 35

Parathyroid hormone-related peptide is the major factor responsible for hypercalcemia of malignancy. There is increasing evidence that parathyroid hormone-related peptide also plays an important role in the growth and differentiation of both neoplastic and non-neoplastic cells. Recently we found that reactive human bile ductules and cholangiocarcinomas, but not normal bile ducts, human hepatocytes nor hepatocellular carcinomas, express parathyroid hormone-related peptide and we speculated that parathyroid hormone-related peptide may function as a growth and differentiation factor for bile ductular epithelial cells. Using a specific polyclonal antibody for immunostaining and a digoxigenin-random prime-labeled probe for in situ hybridization assay, we found that only cell lines with a bile duct phenotype expressed parathyroid hormone-related peptide and its mRNA. HepG2 cells with hepatocellular phenotype (CK19-, CK7-, CK8+, CK18+, albumin+) do not express parathyroid hormone-related peptide. However, A16 (HepG2 derived cell line) expressing bile duct marker CK19, also expressed parathyroid hormone-related peptide, while hepatocyte markers CK8, CK18, CALLA and albumin were negative. In addition, the H1 cell line (adult human hepatocytes immortalized in our laboratory by SV40 DNA transfection, passaged at least 40 times and cultured for 13 months) expressed bile duct marker CK7 and parathyroid hormone-related peptide, while hepatocyte markers CK8, CK18, CALLA and albumin were negative. Previous studies demonstrated that parathyroid hormone-related peptide gene expression in keratinocytes can be modulated by serum, growth factors and cycloheximide although there is a species and cellular specificity.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Parathyroid hormone-related peptide is expressed and rapidly inducible in human liver cell cultures that have a bile duct phenotype. 749 87

The effects of retinoic acid (RA) on the expression of osteoblastic-related cell markers was examined. A marrow stromal osteogenic cell line, MBA-15, was analyzed by Northern blotting for the expression of bone matrix proteins. These cells constitutively express mRNA encoding for procollagen alpha 2 (I), osteonectin, osteopontin, biglycan, and alkaline phosphatase (ALK-P). Gene expression was unchanged in response to RA triggering for 24 hr. Furthermore, cell growth and enzymatic activities of ALK-P and neutral endopeptidase (CD10/NEP) were studied. These parameters were examined in MBA-15 and clonal populations representing different stages of differentiation. The cell's growth rate was unchanged, while ALK-P activity was greatly increased during the culture period under RA treatment in MBA-15 and in the clonal cell lines examined while CD10/NEP activity displayed a different pattern. MBA-15.4, a preosteoblast cell line, exhibited an inhibition in CD10/NEP activity at the beginning of the culture period, reaching basal level with time. This activity was greatly increased over control level in MBA-15.6, a mature stage of osteoblasts. Furthermore, the response of cell lines to various growth factors was tested subsequent to priming the cultures with RA. A synergistic effect was monitored for ALK-P activity in MBA-15.4 and MBA-15.6 cells under rh-bone morphogenic protein (BMP-2) and purified osteogenin (BMP-3), and an antagonist effect was measured when cells were exposed to transforming growth factor beta (TGF beta). Contrarily, BMP-2 and BMP-3 inhibited the CD10/NEP activity that had remained unchanged following priming of the cell with RA. Insulin-like growth factor I (IGF-I) and basic fibroblast growth factors (bFGF) did not affect either ALK-P nor CD10/NEP activities in both cloned cells. Cellular response to bone-seeking hormone, parathyroid hormone (PTH), and prostaglandin E2 (PGE2) was monitored by activation of intracellular cAMP. Treatment with RA caused a dramatic decrease in MBA-15.6 cell responses to PTH and PGE2, but no significant effects could be observed in other clonal lines.
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PMID:Differential effects of retinoic acid and growth factors on osteoblastic markers and CD10/NEP activity in stromal-derived osteoblasts. 752 53


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