UNIPROT:P15088 - FACTA Search

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Query: UNIPROT:P15088 (mast cell)
14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human chymase and rat chymase-1 are mast cell serine proteases involved in angiotensin II (Ang II) formation and degradation, respectively. Previous studies indicate that both these enzymes have similar P1 and P2 preferences, which are the major determinants of specificity. Surprisingly, despite the occurrence of optimal P2 and P1 residues at the Phe8 downward arrow and Tyr4 downward arrow bonds (where downward arrow, indicates the scissile bond in peptide substrates) in Ang I (DRVYIHPFHL), human chymase cleaves the Phe8 downward arrow bond with an approximately 750-fold higher catalytic efficiency (kcat/Km) than the Tyr4 downward arrow bond in Ang II (DRVYIHPF), whereas rat chymase-1 cleaves the Tyr4 downward arrow bond with an approximately 20-fold higher catalytic efficiency than the Phe8 downward arrow bond. Differences in the acyl groups IHPF and DRVY at the Phe8 downward arrow and Tyr4 downward arrow bonds, respectively, are chiefly responsible for the preference of human chymase for the Phe8 downward arrow bond. We show that the IHPF sequence forms an optimal acyl group, primarily through synergistic interactions between neighboring acyl group residues. In contrast to human chymase, rat chymase-1 shows a preference for the Tyr4 downward arrow bond, mainly because of a catalytically productive interaction between the enzyme and the P'1 Ile5. The overall effect of this P'1 Ile interaction on catalytic efficiency, however, is influenced by the structure of the acyl group and that of the other leaving group residues. For human chymase, the P'1 Ile interaction is not productive. Thus, specificity for Ang II formation versus Ang II degradation by these chymases is produced through synergistic interactions between acyl or leaving group residues as well as between the acyl and leaving groups. These observations indicate that nonadditive interactions between the extended substrate binding site of human chymase or rat chymase-1 and the substrate are best explained if the entire binding site is taken as an entity rather than as a collection of distinct subsites.
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PMID:Distinct multisite synergistic interactions determine substrate specificities of human chymase and rat chymase-1 for angiotensin II formation and degradation. 900 43

Sheep mast cell proteinase 1 (SMCP-1), which is abundantly expressed in gastrointestinal but not skin mast cells, was isolated and its substrate specificity was investigated. Peptide substrates, including angiotensin I, substance P, bradykinin and oxidized insulin B chain were hydrolysed at P1 Phe, Leu or Tyr residues, conforming to the known chymotrypsin-like properties of the enzyme. However, SMCP-1 was found to hydrolyse some chromogenic substrates with P1 Lys and Arg residues. The enzyme also demonstrated trypsin-like activity against protein substrates, cleaving BSA at Lys114-Leu115, Lys238-Val239, Lys260-Tyr261 and Lys376-His377. Bovine fibrinogen beta-chain was cleaved at Lys28-Lys29. To ensure homogeneity of the enzyme, the ratio of chymotrypsin-like to trypsin-like activity was observed; it was found to be constant during purification and between different preparations of SMCP-1. Treatment of SMCP-1 with a range of inhibitors decreased chymotrypsin-like and trypsin-like activities by similar extents, supporting the assertion that both activities are the property of a single enzyme. In terms of activity, and by N-terminal amino acid sequencing, SMCP-1 strongly resembles the similarly dual-specific bovine duodenal proteinase, duodenase. It is proposed that SMCP-1 and duodenase represent a new class of ruminant chymases with unusual dual specificities.
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PMID:Sheep mast cell proteinase-1: characterization as a member of a new class of dual-specific ruminant chymases. 903 51

In all mammalian species investigated so far, mast cells and basophils are the only cells that synthesize histamine and express plasma membrane receptors that bind IgE with high affinity (Fc epsilonRI). Human basophils and mast cells derive from distinct precursors that originate in the bone marrow and fetal liver and probably circulate in peripheral blood. There is extensive evidence that mast cells and basophils and their mediators are primary effectors of allergic inflammation. Immunologically activated human basophils release two cytokines: IL-4 and IL-13. Expression of several cytokines has been documented in a number of experimental models of human and rodent mast cells. However, to date few studies have analyzed the mechanisms of gene expression in human Fc epsilonRI+ cells. Some of these studies imply a role for NFAT and GATA family members in the IgE-mediated activation of cytokine gene transcription in basophils and mast cells. Studies of human basophils and mast cells isolated from different anatomic sites have established the different profiles of eicosanoids released by these cells. Recently, the characterization of arachidonic acid pools and the identification of novel enzymes involved in arachidonate remodeling and mobilization clarified in part how eicosanoid productions is regulated in mast cells and basophils. In addition to histamine, human mast cell secretory granules contain the neutral proteases tryptase, chymase and carboxypeptidase that possess several biochemical properties. In particular, tryptase may play a role as a fibrogenic factor and chymase might convert angiotensin I to angiotensin II. Mast cells are present in human heart and in human coronary arteries raising the possibility that local activation of cardiac mast cells might contribute to certain cardiovascular diseases. Recent evidence also suggests that mast cells and basophils can play a role during viral and bacterial infections. It is now evident that in man these two cells not only participate in inflammation associated with allergic disease, but also in chronic and fibrotic disorders affecting several organs and in host defense against bacterial and viral infections.
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PMID:Molecular and cellular biology of mast cells and basophils. 936

A chymotrypsin-like proteinase, designated myonase, was successfully purified to homogeneity from X-chromosome linked muscular dystrophic mouse skeletal muscle by affinity chromatography on agarose conjugated with lima bean trypsin inhibitor as ligand. The molecular mass of the purified myonase was determined to be 26 kDa by SDS-PAGE and to be 25,187 Da by mass spectrometry. The native enzyme is a single chain molecule and a monomeric protein without sugar side-chains. The nucleotide sequence of myonase mRNA is similar to mouse mast cell proteinase 4 (MMCP-4) cDNA. This is the first report of a native enzyme whose amino acid sequence closely corresponds to MMCP-4 cDNA. Myonase has chymotrypsin-like activities and hydrolyzes the amide bonds of synthetic substrates having Tyr and Phe residues at the P1 position. Myonase is most active at pH 9 and at high concentration of salts. Myonase preferentially hydrolyzes the Tyr4-Ile5 bond of angiotensin I and the Phe20-Ala21 bond of amyloid beta-protein, and it is less active towards the Phe8-His9 bond of angiotensin I and the Phe4-Ala5 and Tyr10-Glu11 bonds of amyloid beta-protein. Myonase is completely inhibited by such serine proteinase inhibitors as chymostatin, diisopropylfluorophosphate and phenylmethylsulfonyl fluoride, but not by p-tosyl-L-phenylalanine chloromethyl ketone, p-tosyl-L-lysine chloromethyl ketone, pepstatin, E-64, EDTA, and o-phenanthroline. It is also inhibited by lima bean trypsin inhibitor, soy bean trypsin inhibitor, and human plasma alpha1-antichymotrysin. These properties match those of chymase, but unlike chymase, myonase does not interact with heparin in the regulation of its activity. Myonase was immunohistochemically localized in myocytes, but not in mast cells.
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PMID:Purification and characterization of myonase from X-chromosome linked muscular dystrophic mouse skeletal muscle. 953 57

The enzymatic pathways for local angiotensin II (Ang II) formation in the heart have been studied both in vivo and in vitro, but the results of these experiments have been discrepant. Thus, the experiments in vivo with intact hearts, both in humans and in animal models, have unequivocally demonstrated that the major Ang II-forming enzyme is angiotensin-converting enzyme (ACE). In contrast, the experiments in vitro with both human or animal heart preparations, have unequivocally demonstrated that the major Ang II-forming enzyme is chymase, a mast cell-derived chymotrypsin-like serine protease. The in vitro approach, however, seems to involve several pitfalls, which tend to overestimate the contribution of chymase as compared to that of ACE. It seems evident that in vivo the chymase-mediated Ang II formation is subjected to local inhibition, a fact that has been overlooked in most of the studies performed in vitro. Accordingly, human chymase, even in its natural form as a protease-proteoglycan complex, is highly sensitive to the protease inhibitors naturally present in the interstitial fluid (IF). We found that if human heart tissue preparations are incubated in vitro in the presence of IF, the chymase-mediated Ang II formation is almost totally suppressed. As the heart interstitium is constantly bathed by IF with its protease inhibitors in concentrations sufficiently high to ensure efficient inhibition of this enzyme, the protease inhibitor-mediated suppression of chymase should also be effective in vivo. Thus, the local production of Ang II in the human heart appears to be regulated by ACE rather than by chymase.
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PMID:Angiotensin II formation in the human heart: an ACE or non-ACE-mediated pathway? 980 Aug 78

Angiotensin II (Ang II) has powerful modulatory actions on cardiovascular function that are mediated by specific receptors located on neurons within the hypothalamus and brain stem. Incubation of neuronal cocultures of rat hypothalamus and brain stem with Ang II elicits an Ang II type 1 (AT1) receptor-mediated inhibition of total outward K+ current that contributes to an increase in neuronal firing rate. However, the exact K+ conductance(s) that is inhibited by Ang II are not established. Pharmacological manipulation of total neuronal outward K+ current revealed a component of K+ current sensitive to quinine, tetraethylammonium, and 4-aminopyridine, with IC50 values of 21.7 micromol/L, 1.49 mmol/L, and 890 micromol/L, respectively, and insensitive to alpha-dendrotoxin (100 to 500 nmol/L), charybdotoxin (100 to 500 nmol/L), and mast cell degranulating peptide (1 micromol/L). Collectively, these data suggest the presence of Kv2.2 and Kv3.1b. Biophysical examination of the quinine-sensitive neuronal K+ current demonstrated a macroscopic conductance with similar biophysical properties to those of Kv2.2 and Kv3.1b. Ang II (100 nmol/L), in the presence of the AT2 receptor blocker PD123,319, elicited an inhibition of neuronal K+ current that was abolished by quinine (50 micromol/L). Reverse transcriptase-polymerase chain reaction analysis confirmed the presence of Kv2.2 and Kv3.1b mRNA in these neurons. However, Western blot analyses demonstrated that only Kv2.2 protein was present. Coexpression of Kv2.2 and the AT1 receptor in Xenopus oocytes demonstrated an Ang II-induced inhibition of Kv2.2 current. Therefore, these data suggest that inhibition of Kv2.2 contributes to the AT1 receptor-mediated reduction of neuronal K+ current and subsequently to the modulation of cardiovascular function.
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PMID:Angiotensin II type 1 receptor-mediated inhibition of K+ channel subunit kv2.2 in brain stem and hypothalamic neurons. 1002 10

The effects of cold-restraint stress, repeated over 3 days, and treatment of rats with vasoactive intestinal peptide (VIP) on the contractile responses of isolated aorta to vasoconstrictors, and on aortic adventitial mast cells were investigated. Stress significantly reduced the contractile response of rat aorta smooth muscle to norepinephrine (NE), angiotensin II (Ang II) and vasopressin (VP). Decreased sensitivity to NE, Ang II and VP may result from decreased receptor density, and affinity or reduced effector efficacy. Stress induced degranulation, decreased the number and changed the granular content of mast cells; all degranulated mast cells were stained with alcian blue, and the percentage of safranin staining cells was decreased. Given prior to stress, VIP reversed the reduced contractile responses and sensitivity of aorta to NE and Ang II but had no effect on VP subsensitivity. VIP also inhibited stress-induced degranulation of mast cells, and after VIP only alcian blue-stained mast cells were seen. When VIP was given to non-stressed rats, the contractile response of the aorta to NE, but not Ang II or VP, was increased compared with control. Mast cell count was decreased in the adventitia of non-stressed VIP treated rats. The results indicate that stress decreases the heparin content of mast cells and VIP has an additive effect. In conclusion, VIP modulates both stress-induced mast cell activity and reduced sensitivity of aorta smooth muscle to NE and Ang II. It can be suggested that VIP may moderate some effects of stress on vascular pathophysiology.
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PMID:The effect of stress and in vivo vasoactive intestinal peptide (VIP) treatment on the response of isolated rat aorta to norepinephrine, angiotensin II and vasopressin, and adventitial mast cells. 1134 95

We investigated the degradation of angiotensin I (Ang I) by guinea pig aqueous humor at physiological pH (pH 7.4) and assessed the activity of responsible enzymes using various enzyme inhibitors. The aqueous humor was incubated with Ang I in the presence or absence of an enzyme inhibitor at 37 degrees C for the appropriate time period. The resulting peptides were analyzed by a Beckman HPLC system with a Waters microBondapak C18 analytical column using a 30-min increasing linear gradient of 10 to 40% acetonitrile containing 0.05% trifluoroacetic acid (TFA) and H2O containing 0.05% TFA at a flow rate of 1 mL/min. Detection was done by absorbance at 214 nm. Angiotensin II (Ang II) was a major product (39.3+/-4.10 nmol x h(-1) mL(-1), n = 5) of Ang I hydrolysis. Traces of angiotensin 1-9, angiotensin IV, and angiotensin 1-7 were also produced. Chymostatin (0.05 mmol/L), EDTA (1 mmol/L), enalaprilat (0.1 mmol/L), and ebelacton B (0.01 mmol/L) inhibited generation of Ang II from Ang I by guinea pig aqueous humor by 89+/-4.6, 56+/-7.6, 33+/-5.1, 20+/-6.5%, respectively. Our findings indicate that guinea pig aqueous humor contains several enzymes that can form Ang II. The chymostatin-sensitive type of enzyme was the most active one found in guinea pig aqueous humor. Angiotensin I converting enzyme, carboxypeptidase A, and deamidase may also contribute to angiotensin II formation in guinea pig ocular fluid.
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PMID:Metabolism of angiotensin I by guinea pig aqueous humor. 1147 97

Angiotensin-converting enzyme (ACE) has a critical role in cardiovascular function by cleaving the carboxy terminal His-Leu dipeptide from angiotensin I to produce a potent vasopressor octapeptide, angiotensin II. Inhibitors of ACE are a first line of therapy for hypertension, heart failure, myocardial infarction and diabetic nephropathy. Notably, these inhibitors were developed without knowledge of the structure of human ACE, but were instead designed on the basis of an assumed mechanistic homology with carboxypeptidase A. Here we present the X-ray structure of human testicular ACE and its complex with one of the most widely used inhibitors, lisinopril (N2-[(S)-1-carboxy-3-phenylpropyl]-L-lysyl-L-proline; also known as Prinivil or Zestril), at 2.0 A resolution. Analysis of the three-dimensional structure of ACE shows that it bears little similarity to that of carboxypeptidase A, but instead resembles neurolysin and Pyrococcus furiosus carboxypeptidase--zinc metallopeptidases with no detectable sequence similarity to ACE. The structure provides an opportunity to design domain-selective ACE inhibitors that may exhibit new pharmacological profiles.
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PMID:Crystal structure of the human angiotensin-converting enzyme-lisinopril complex. 1254 Aug 54

In this study we demonstrate that carboxypeptidase A (CPA)-like enzyme is expressed in rat kidney. The major metabolites of angiotensin (Ang) I by the rat renal mesangial cell extract at 37 degrees C, pH 7.4, were Ang 1-9 and Ang II. Quinaprilat did not influence the formation of Ang 1-9, but it inhibited formation of Ang II. The formation of Ang 1-9 was inhibited by potato carboxypeptidase inhibitor, 1,10-phenanthroline or EDTA. Lowering the pH from 7.4 to 4.0 also inhibited the formation of this nonapeptide. These findings suggest that a metallocarboxypeptidase is responsible for Ang 1-9 production. Using monoclonal antibodies to CPA, Western blot showed the presence of CPA-like enzyme in the extracts prepared from the mesangial cells or kidney cortex of the rat. Immunohistochemistry showed that CPA-like enzyme is localized in the mesangial glomerular cells and adventitia of kidney blood vessels, whereas it was absent in the renal tubules. Our data suggest that a CPA-like enzyme could be added to a repertoire of enzymes present in the rat mesangial cells and adventitia of renal blood vessels.
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PMID:Localization of carboxypeptidase A-like enzyme in rat kidney. 1461 96

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