Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Thrombin-mediated down-regulation of endothelin (ET) receptors was studied in rat glomerular mesangial cells. Overnight incubation of mesangial cells with thrombin (10 nM) resulted in a significant decrease (67%) in the number of ET receptors, with no change in affinity. Northern analysis of the mRNA from these cells showed a corresponding decrease in the ETA receptor message. Such a decrease in ET receptors could result from an increase in ET levels caused by an increase in synthesis and/or a decrease in degradation. It has been previously reported that thrombin stimulates ET production in endothelial and mesangial cells. Because ET is known to be degraded by neutral endopeptidase (NEP), which is present at high levels in the kidney, the potential effects of thrombin on NEP activity were evaluated. There was a decrease of NEP activity in mesangial cells at 16 and 24 hr after treatment with 10 nM thrombin. This effect was specific for thrombin, because NEP activity was not altered after treatment with thrombin in the presence of hirudin, an inhibitor of thrombin activity. The thrombin-mediated decrease in NEP activity correlated with a decrease in NEP protein and mRNA levels, as determined by Western and Northern analyses, respectively. To determine whether the thrombin-mediated decrease in ET receptors had a functional corollary, ET-1-stimulated intracellular calcium mobilization was measured. Overnight incubation with 10 nM thrombin resulted in a significant inhibition of ET-stimulated intracellular calcium mobilization. This effect was specific for ET, because thrombin pretreatment did not affect vasopressin-stimulated intracellular calcium mobilization in mesangial cells. These results indicate that the thrombin-mediated down-regulation of ET receptors is due, in part, to a thrombin-stimulated increase in ET resulting from the down-regulation of NEP and the reported increase in ET synthesis. In addition, pretreatment of mesangial cells with ET-1 caused a significant decrease (85%) in ET receptor number and ET-1-mediated intracellular calcium release (84%), without affecting vasopressin- or thrombin-mediated responses.
Mol Pharmacol 1995 Jun
PMID:Thrombin-mediated down-regulation of endothelin receptors in mesangial cells coincides with the down-regulation of neutral endopeptidase activity. 760 55

Alveolar macrophages protect the lungs against noxious agents. Proteases and peptidases are essential for this defense and many metabolic activities. Human alveolar macrophages were evaluated for the presence of six important peptidases. Deamidase, a serine peptidase identical with the lysosomal protective protein and possibly with cathepsin A, had high specific activity in alveolar macrophages and is also present in cultured mouse J774A.1 and human U937 cells, used for the sake of comparison. In fractionated J774A cells, most of the deamidase activity was in the lysosomal fraction and in the final supernatant. Deamidase in human alveolar macrophages, obtained by bronchoalveolar lavage from 23 patients, cleaved dansyl-Phe-Leu-Arg at a rate of 2.26 mumol/h/mg protein and hydrolyzed the chemotactic peptide N-f-Met-Leu-Phe even faster, at a rate of 53.1 mumol/h/mg protein, the highest activity for this enzyme with any of the cells we tested. Rabbit antiserum, elicited with the recombinant partial sequence of the enzyme, immunoprecipitated 77-88% of the macrophage deamidase. In immunocytochemistry, this antiserum localized deamidase within the human macrophages. The enzyme was inhibited by diisopropylfluorophosphate (DFP; 1 mM) and by ebelactone B (10 microM), noncompetitively. The mRNA of deamidase was detected in mouse macrophages by Northern blot; the two protein chains of deamidase were shown in human macrophages by Western blot. In addition, two other serine peptidases were also highly active in macrophages: dipeptidyl peptidase IV (1.38 mumol/h/mg protein) and prolylcarboxypeptidase (0.72 mumol/h/mg protein). The activity of plasma membrane zinc metallopeptidases, neutral endopeptidase 24.11 and carboxypeptidase M, in contrast, was low or absent (angiotensin I converting enzyme; kininase II).(ABSTRACT TRUNCATED AT 250 WORDS)
Am J Respir Cell Mol Biol 1995 Aug
PMID:Plasma membrane-bound and lysosomal peptidases in human alveolar macrophages. 762 87

Epstein-Barr virus (EBV) negative and EBV carrying Burkitt lymphoma (BL) lines that remain phenotypically similar to the in vivo tumor cells (operationally defined group I BLs) express high levels of CD10 and CD77, and lack immunoblastic markers such as CD23 and CD39, and the cell adhesion molecules CD11a, CD18, CD54 and CD58. This cell phenotype is associated with poor stimulatory capacity in allogeneic mixed lymphocytes cultures (MLC) [Avila-Carino et al. Int. J. Cancer 40, 691-697 (1987)] EBV carrying BL lines tend to drift spontaneously towards an immunoblastic phenotype in parallel with up-regulation of six EBV-encoded nuclear antigens (EBNA-2 to -6) and two membrane proteins (LMP-1 and -2). These viral antigens are characteristically expressed in all EBV transformed lymphoblastoid cell lines (LCLs) of normal B cell origin and can be induced in group I BL lines by treatment with the DNA demethylating agent 5-azacytidine (5-azaC) [Masucci et al. J. Virol. 65, 1558-1567 (1989)]. We have now studied the effect of 5-azaC on the induction of allogenic T cell proliferation by three EBV negative (Ramos, BL28 and BL41) and four EBV carrying BL lines (Rael, Eli, Chep and Mutu) which stably express a group I phenotype. Pre-treatment with 4-15 microM 5-azaC had no effect on the EBV negative cells but increased the stimulatory capacity of all four EBV carrying lines. LMP-1 was the only viral antigen regularly induced suggesting that its expression may be required for the increase of allostimulation. This was corroborated by the observation that LMP-1 transfection increased 35-70-fold the stimulatory capacity of Rael cells. The cell adhesion molecule CD54 was the only cellular marker selectively up-regulated in all cell lines with increased stimulatory capacity.
Mol Immunol 1993 Apr
PMID:Selective induction of allostimulatory capacity after 5-azaC treatment of EBV carrying but not EBV negative Burkitt lymphoma cell lines. 768 32

We are interested in the mechanisms of ozone-induced lung effects after short-term exposure and the relationship with subsequent pulmonary inflammation and disease. Our hypothesis is that ozone, as a powerful oxidant, will diminish the activity of neutral endopeptidase (NEP) in the airways of humans with resulting increased concentrations of neuropeptides such as substance P (SP). We have exposed seven (two women, five men) healthy, nonsmoking individuals (22 to 30 yr of age) to filtered air and ozone (0.25 ppm) for 1 h in an environmental chamber during heavy exercise. Bronchoscopy with airway lavage (AL) and bronchoalveolar lavage (BAL) was performed immediately after ozone exposure. The lavage samples were analyzed by enzyme immunoassay for SP and 8-epi-prostaglandin F2 alpha (8-epi-PGF2 alpha) (a marker for oxidative free radical reaction) and by radioimmunoassay for complement fragments. FEV1 had declined 12.4 +/- 1.9% (mean +/- SEM) as a result of ozone exposure. The AL concentration for SP and 8-epi-PGF2 alpha and BAL concentration of C3a after ozone exposure were significantly higher than after the filtered air exposure (P < 0.05). There was a significant correlation between SP and 8-epi-PGF2 alpha concentrations in the AL fluid (r2 = 0.89 and P < 0.05). There were no changes in C5a in either compartment or any of the mediators in the plasma samples. These results extend previous results from animal studies suggesting that ozone's mechanism of action is through an oxidative reaction resulting in a decreased activity of NEP in the airways with a subsequent increase in the concentration and activity of SP.
Am J Respir Cell Mol Biol 1993 Nov
PMID:Ozone-induced increases in substance P and 8-epi-prostaglandin F2 alpha in the airways of human subjects. 769 98

Biochemical studies of pollen proteins have been focused, primarily, in investigating their roles as allergens. These molecules, some of which have enzymatic activity, act as antigens and initiate the production of IgE antibodies, leading to allergic and/or asthmatic responses. Included in this mixture of proteins are proteinases which, although they may or may not be allergenic, could still be involved in airway dysfunction. We have isolated an arginine-specific endopeptidase to homogeneity from mesquite (Prosopis velutina) pollen, a known wind-borne allergen, which has a molecular mass near 84 kDa by NaDodSO4-gel electrophoresis, a pH optimum in the neutral to alkaline range, and a requirement for Ca2+ for stabilization. The enzyme is inhibited by diisopropyl fluorophosphate (DFP) and N-p-tosyl-L-lysine chloromethylketone but not by N-p-tosyl-L-phenylalanine chloromethylketone, EDTA, or iodoacetamide. It was also not inhibited by human plasma proteinase inhibitors nor several other naturally occurring plant and animal inhibitors. Cleavage by the endopeptidase was primarily on the carboxy-terminal side of arginine residues in peptides, whereas proteins such as kallikrein and prothrombin were only activated and/or degraded extremely slowly. Several bioactive peptides that may be involved in maintaining normal lung function were readily fragmented, including angiotensin II, a vasoconstrictor, and atrial natriuretic peptide, a modulator of vascular permeability, both of which were rapidly cleaved at low enzyme:substrate molar ratios. Thus, the pollen endopeptidase could be involved in exacerbating the development of asthma by inactivating bioactive peptides that have ameliorating effects in maintaining lung airway homeostasis.
Am J Respir Cell Mol Biol 1995 Apr
PMID:Isolation and properties of an angiotensin II-cleaving peptidase from mesquite pollen. 769 24

We have identified neutral proteolytic activities that convert big endothelin-1 (big ET-1) to ET-1 in guinea-pig lung membrane fraction. The active proteins have been solubilized and two distinct enzymes have been purified by combinations of sequential column chromatographies. One purified enzyme was a metalloenzyme based upon its sensitivity to chelating agent with a molecular mass of 108 kDa on SDS-PAGE. Another enzyme was also a metalloenzyme with a molecular mass of 162 kDa. Further investigations revealed that the 108 kDa enzyme was inhibited by phosphoramidon and also by thiorphan, and produced ET-1 with the Km value of 5.7 microM for big ET-1. The 162 kDa enzyme was also inhibited by phosphoramidon, but neither by thiorphan nor by captopril, and quantitatively produced ET-1 from big ET-1 with a Km value of 2.1 microM. These results indicate that the 108 kDa enzyme probably seems to be a neutral-endopeptidase (EC 3. 4. 24. 11.) or similar one, but the 162 kDa enzyme is a unique metalloprotease that converts big ET-1 to ET-1.
Biochem Mol Biol Int 1994 Dec
PMID:Endothelin converting enzymes in guinea-pig lung membrane fractions: purifications and characterizations. 769 95

We have purified initiatorin, a prostatic endopeptidase that initiates the protein-arginine degradation cascade in the spermatophore of Bombyx mori. Purification of the enzyme from spermatophores was monitored by measuring BAEE (N alpha-benzoyl-L-arginine-ethyl ester) hydrolyzing activity. Spermatophores were used as a source for this enzyme. Of several isoforms the major form (MW, 29 kDa) was purified over 200-fold. The N-terminal sequence of initiatorin showed strong homology with those of serine-type of endopeptidases.
Insect Biochem Mol Biol 1994 Dec
PMID:Purification and partial amino acid sequence of initiatorin, a prostatic endopeptidase of the silkworm, Bombyx mori. 770 88

Schwann cells in culture divide in response to defined mitogens such as PDGF and glial growth factor (GGF), but proliferation is greatly enhanced if agents such as forskolin, which increases Schwann cell intracellular cAMP, are added at the same time as PDGF or GGF (Davis, J. B., and P. Stroobant. 1990. J. Cell Biol. 110:1353-1360). The effect of forskolin is probably due to an increase in numbers of PDGF receptors (Weinmaster, G., and G. Lemke. 1990. EMBO (Eur. Mol. Biol. Organ.) J. 9:915-920. Neuropeptides and beta-adrenergic agonists have been reported to have no effect on potentiating the mitogenic response of either PDGF or GGF. We show that the neuropeptide calcitonin gene-related peptide (CGRP) increases Schwann cell cAMP levels, but the cells rapidly desensitize. We therefore stimulated the cells in pulsatile fashion to partly overcome the effects of desensitization and show that CGRP can synergize with PDGF to stimulate Schwann cell proliferation, and that CGRP is as effective as forskolin in the pulsatile regime. CGRP is a good substrate for the neutral endopeptidase 24.11. Schwann cells in vivo have this protease on their surface, so the action of CGRP could be terminated by this enzyme and desensitization prevented. We therefore suggest that CGRP may play an important role in stimulating Schwann cell proliferation by regulating the response of mitogenic factors such as PDGF.
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PMID:Calcitonin gene-related peptide promotes Schwann cell proliferation. 773 Apr 12

The selection of therapeutic agents for clinical development is principally based upon pharmacodynamic and pharmacokinetic criteria. Although many compounds are routinely tested in pharmacologic assays, direct pharmacokinetic assessment is difficult and not applicable to a large number of agents. Therefore, we have developed a rapid indirect method based on enzyme inhibition for determining the unbound concentration of NEP 24.11 inhibitors in rat plasma. In the present study, drug levels of compounds from three different classes of NEP 24.11 inhibitors: mercaptoalkyl, carboxyalkyl and aminophosphonates were compared. Studies were carried out in conscious, unrestrained rats. Arterial blood samples were obtained 10, 30, 60, 120, and 240 min after drug administration at 10 mg/kg i.v. or 30 mg/kg p.o. The blood was collected in EDTA and plasma prepared immediately. Protein bound NEP inhibitor was separated from plasma by centrifugation through an ultrafiltration membrane. Following acidification and serial dilution, the concentration of unbound inhibitor was determined in the plasma ultrafiltrate using the in vitro assay for NEP 24.11 inhibition. The results of this study indicated that the mercaptoalkyl inhibitor thiorphan was cleared rapidly from plasma, whereas, the plasma concentrations of the carboxyalkyl inhibitor CGS 23880A (UK-69,578), and the plasma concentrations of the aminophosphonate, CGS 24128, were maintained at high levels for at least 4 hours. Furthermore, the ratio of the NEP inhibitor concentration/IC50 value correlated well with the pharmacologic activity of these compounds.
Res Commun Mol Pathol Pharmacol 1995 Feb
PMID:A rapid indirect method to determine the plasma concentrations of neutral endopeptidase inhibitors. 774 58

The three-dimensional structure of the calcium-free form of Bacillus licheniformis alpha-amylase (BLA) has been determined by multiple isomorphous replacement in a crystal of space group P4(3)2(1)2 (a = b = 119.6 A, c = 85.4 A). The structure was refined using restrained crystallographic refinement to an R-factor of 0.177 for 28,147 independent reflections with intensities FObs > 0 at 2.2 A resolution, with root mean square deviations of 0.008 A and 1.4 degrees from ideal bond lengths and bond angles, respectively. The final model contains 469 residue, 237 water molecules, and one chloride ion. The segment between Trp182 and Asn192 could not be located in the electron density, nor could the N and C termini. Cleavage of the calcium-free form of BLA was observed after Glu189, due to a Glu-C endopeptidase present in trace amounts in the preparation. BLA did not crystallize without this cleavage under the conditions applied. BLA exhibits the characteristic overall topological fold observed for other alpha-amylases and related amylolytic enzymes: a central domain A containing an alpha/beta-barrel with a large protrusion between beta-strand 3 and alpha-helix 3 (domain B) and a C-terminal greek key motif (domain C). Unlike in the other enzymes, domain B possesses a beta-sheet made up of six loosely connected, twisted beta-strands forming a kind of a barrel with a large hole in the interior. Topological comparisons to TAKA-amylase, pig pancreatic alpha-amylase and cyclodextrin glycosyltransferase reveal a very high structural equivalence for large portions of the proteins and an exceptionally pronounced structural similarity for calcium binding, chloride binding and the active site. None of the theories proposed to explain the enhanced thermostability of BLA showed a satisfactory correlation with the three-dimensional structure. Instead, sequence comparisons to the less thermostable bacterial alpha-amylase from Bacillus amyloliquefaciens (BAA) indicate that some ionic interactions present in BLA, but which cannot be formed in BAA, might be responsible for the enhanced thermostability of BLA.
J Mol Biol 1995 Mar 03
PMID:Crystal structure of calcium-depleted Bacillus licheniformis alpha-amylase at 2.2 A resolution. 787 75


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