Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A gene encoding human chymase was cloned and sequenced. The protein-coding exons reveal a preproenzyme with a 19-amino acid signal peptide, an acidic 2-amino acid propeptide, and a 226-amino acid catalytic domain. The mature enzyme is predicted to be cationic (net charge of +13) and to be modified by N-glycosylation at two sites. The amino acid sequence is identical to the 35 residues of NH2-terminal amino acid sequence reported for human skin chymase and is identical to 29 of 31 residues of NH2-terminal and internal amino acid sequence reported for human heart chymase. The full predicted sequence of the catalytic domain reveals a high level of sequence identity to dog mast cell chymase (83%) and a lower level of identity to the sequences of rodent chymases (58-62%). In the phase and placement of introns, the organization of this human chymase gene is similar to that of several other granule-associated leukocyte serine proteases, including rat chymase II, lymphocyte granzymes, and neutrophil cathespin G and elastase. However, the gene organization differs from that of mast cell tryptase, providing additional evidence that the major mast cell serine proteases are separated by substantial evolutionary distance. Amplification of chymase gene-specific fragments from hamster/human hybrid cell line DNA suggests localization of the chymase gene to human chromosome 14. High stringency hybridization of chymase DNA to a human genomic DNA blot suggests the possibility of more than one human chymase gene. Evidence that the chymase gene is expressed in human tissues was obtained by the amplification of chymase-specific DNA from skin and placental cDNA libraries.
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PMID:Structure, chromosomal assignment, and deduced amino acid sequence of a human gene for mast cell chymase. 207 82

Tryptase, a neutral endoprotease, is secreted by activated mast cells in human tissues. Tryptase levels in various body fluids have been used as an indicator of mast cell activation. The authors determined tryptase levels in unstimulated tears collected from the following groups of patients: (1) normal control, (2) nonallergic ocular inflammation, (3) asymptomatic seasonal allergic conjunctivitis, (4) symptomatic seasonal allergic conjunctivitis, (5) vernal conjunctivitis, and (6) contact lens-associated giant papillary conjunctivitis. They also assessed the release of tryptase into the tear fluid after provoking the conjunctiva with (7) allergens, (8) compound 48/80, and (9) rubbing. Tryptase levels were elevated in tears of patients with active ocular allergy and also increased after provoking the conjunctiva with allergens in atopic subjects and with compound 48/80 and rubbing in nonatopic subjects. Tryptase levels in tear fluid may prove useful as a clinical indicator of mast cell involvement in ocular allergic disorders. In provocation experiments, tryptase levels may be used to evaluate and compare different mast cell stabilizing agents.
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PMID:The level of tryptase in human tears. An indicator of activation of conjunctival mast cells. 208 98

The neutral protease tryptase has been isolated from a human mast cell line, HMC-1. The HMC-1 line was established from the peripheral blood of a patient with mast cell leukemia and maintained as continuously proliferating clones in vitro and as solid mast cell tumors in nude mice. HMC-1-derived tryptase was purified by sequential chromatography on Dowex 1, DEAE 5 PW, and heparin-agarose. Purified tryptase has an apparent molecular weight of 150,000, as determined by molecular sieve HPLC, but migrates as a doublet of bands of 32/35,000 on SDS-PAGE gels. Maximal enzymatic activity was observed at pH 8.5. Cleavage of tosyl-L-arginine methyl ester by purified tryptase was inhibited by dansyl-L-glutamyl-glycyl-L-arginine chloromethyl ketone 2 HCl, HgCl2, tosyl-L-lysine chloromethyl ketone, leupeptin, and PMSF but not by benzamidine, aprotinin, tosyl-L-phenyl-alanine chloromethyl ketone, soybean trypsin inhibitor, human plasma, ovomucoid inhibitor, or lima bean trypsin inhibitor. Microsequencing of purified tryptase yielded an amino terminal sequence that was identical to that previously reported for human pituitary-derived tryptase.
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PMID:Purification of tryptase from a human mast cell line. 211 May 91

We have previously characterized dog mastocytoma cells propagated in nude mice. We have established two of these lines (C1 and C2) in continuous culture. Freshly disaggregated mastocytoma cells were cultured in Dulbecco's modified Eagle's medium (DME)-H16 mixed with 50% Ham's F12 and supplemented with histidine and 5% allergic dog serum (ADS). Cells were fed every 3 d and passaged weekly. Growth was assessed by cell count. Cell growth was best supported by culture in 5% ADS. C1 cells grow in suspension in ADS and have been passaged 55 times with a doubling time of 37.4 +/- 18.7 h (mean +/- 1 SD; n = 15). C2 cells adhere to tissue culture plastic in ADS and have been passaged 26 times with a doubling time of 49.3 +/- 12.5 h (n = 13). Morphologic and functional characteristics are unchanged from those described in cells propagated in nude mice. Histamine content for C1 is 0.46 +/- 0.18 pg/cell (n = 12) and 0.07 +/- 0.04 pg/cell (n = 6) for C2. Both lines contain the neutral protease tryptase and C2 contains chymase. Calcium ionophore A23187 or ragweed antigen caused concentration-dependent histamine release from both cell lines. C1 and C2 generate prostaglandin D2 in response to A23187. We conclude that dog mastocytoma cells can be established in continuous culture, thus providing a system for studying mast cell biology, including growth and development.
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PMID:Establishment of two dog mastocytoma cell lines in continuous culture. 212 Nov 70

Basal and stimulated changes in ion transport in vitro were examined in jejunal mucosa from rats during inflammation produced after infection with the nematode Nippostrongylus brasiliensis. The gut was acutely inflamed at days 7 and 10 when net secretion of Na+ and Cl- ions was evident. Serum levels of rat mast cell protease II were elevated, providing evidence for mast cell activation. In addition, the magnitude of the short-circuit current responses to electrical transmural stimulation of enteric nerves (but not to histamine in the presence of neural blockade) were significantly reduced (p less than 0.01) to 17%-33% of control values, suggesting abnormalities of mucosal nerves. Following worm expulsion, serum levels of rat mast cell protease II and ion transport returned to normal. However, mastocytosis was apparent in gut mucosa and parasite antigen stimulated net secretion. In the absence of antigen, short-circuit current responses to nerve stimulation were increased (to 122% of controls; p less than 0.05). These findings suggest that changes in mast cells and enteric nerves occur during inflammation in this model and implicate neural and mast cell interactions with the epithelium in producing the ion-transport abnormalities.
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PMID:Ion transport abnormalities in inflamed rat jejunum. Involvement of mast cells and nerves. 215 98

The catalytic activity of human tryptase, a mast cell neutral endoprotease, is expressed when the enzyme is in its tetrameric form, but is lost under physiologic conditions concomitant with a quaternary structural alteration involving conversion to a monomeric form. The associated changes in the CD spectra noted in the current study indicate accompanying alterations in the secondary structure of the protein. In particular, the progressive disappearance of the negative minimum centered at 228 nm suggests an effect on beta-sheet structure, which may be important for monomer-monomer interaction and/or stabilization of catalytic activity. Dextran sulfate, like heparin, stabilizes the catalytic activity and quaternary structure of tryptase and also maintains the native secondary structure of the enzyme at and beyond a temperature of 40 degrees C. Dextran sulfate-stabilized tryptase therefore was used as an immunogen to which were produced three murine mAb (B2, C11, and G4) recognizing the catalytically active form of the enzyme. Inactive tryptase bound to plastic microtiter wells was not recognized by any of the newly made antibodies, whereas inactive tryptase in solution was recognized by G4, which when biotinylated, could be used as a detector antibody in a sandwich ELISA for tryptase. Each of the newly made mAb recognized the catalytically active form of tryptase. Thus, alterations in epitopes, perhaps reflecting tertiary structural alterations as well as changes in secondary and quaternary conformations, occur with tryptase inactivation. A pragmatic result of these newly generated antibodies is the affinity purification to homogeneity of active tryptase by sequential chromatography with B2 coupled to CH-Sepharose and heparin-agarose. Tryptase purified by this technique had a specific activity with p-tosyl-L-arginine methyl ester of 117 +/- 9 U/mg and had 3.9 +/- 0.3 active sites per molecule of active enzyme (134,000 m.w.) as titrated with p-nitrophenyl-p'-guanidinobenzoate. The spectral and immunologic data in the current study are consistent with concerted conformational alterations in the secondary and tertiary as well as quaternary structures of tryptase associated with loss of catalytic activity. Failure to reverse any of these alterations with dextran sulfate suggests that the pathway of tetramer assembly in vivo is more complicated than simple subunit association.
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PMID:Immunologic and physicochemical evidence for conformational changes occurring on conversion of human mast cell tryptase from active tetramer to inactive monomer. Production of monoclonal antibodies recognizing active tryptase. 217 9

A sensitive and specific enzyme-linked immunosorbent assay (ELISA) was developed for mouse intestinal mast cell proteinase (IMCP). Specificity was demonstrated by the absence of immunoreactivity with extracts of isolated serosal mast cells (SMC), or with high concentrations (50 micrograms/ml) of the antigenically similar rat mast cell proteinases I or II. The small and large intestines in normal mice were the major sources of IMCP, there being little or no IMCP in non-mucosal tissues. Concentrations of IMCP in normal (non-parasitized) mice were low, but were increased 100-1000-fold intestines of mice infected 10 days earlier with Trichinella spiralis. The kinetic response of secreted IMCP into the blood of mice following infection with T. spiralis was also studied. Systemic release of IMCP coincided with the immune expulsion of adult worms from the intestine, and peak concentrations (9.45 micrograms/ml IMCP) occurred 9 days after infection. The tissue distribution of IMCP, its secretion into blood, and its enteric accumulation during parasite infection, are consistent with a mucosal mast cell (MMC) source for IMCP. The results are discussed in the context of similar findings for rat mast cell proteinase II.
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PMID:Distribution of intestinal mast cell proteinase in blood and tissues of normal and Trichinella-infected mice. 217 29

The effect of chronic dietary antigen challenge on the intestine was examined in sensitized rats. Three groups of Hooded-Lister rats were studied: animals sensitized to egg albumin; sham-sensitized animals; and unmanipulated controls. In sensitized rats, serum immunoglobulin E titers to egg albumin were greater than or equal to 1:64, whereas control and pair-fed rats showed no response. Sensitized rats received egg albumin 1 mg/ml in drinking water and rat chow ad libitum. Pair-fed animals also received egg albumin but were pair-fed with sensitized animals. Controls received water and rat chow ad libitum. Chronic antigen challenge resulted in reduced food intake and weight gain in sensitized animals. When the rats were killed after 9 days of antigen exposure, proximal intestine from experimental animals showed decreased disaccharidase activity, brush-border microvillus surface, area, and villus height. Crypt depth and enterocyte migration rate were increased. Mucosal mast cell involvement was suggested by mast cell proliferation, evidence of mast cell degranulation, and increased serum rat mast cell protease II levels. At the time of death, only sensitized jejunum demonstrated an increase in short-circuit current in Ussing chambers in response to antigen challenge. The findings indicate that chronic antigen exposure leads to intestinal injury, reduced food intake, and diminished weight gain.
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PMID:Intestinal anaphylaxis in the rat. Effect of chronic antigen exposure. 218 52

Three different cDNAs and a gene encoding human skin mast cell tryptase have been cloned and sequenced in their entirety. The deduced amino acid sequences reveal a 30-amino acid prepropeptide followed by a 245-amino acid catalytic domain. The C-terminal undecapeptide of the human preprosequence is identical in dog tryptase and appears to be part of a prosequence unique among serine proteases. The differences among the three human tryptase catalytic domains include the loss of a consensus N-glycosylation site in one cDNA, which may explain some of the heterogeneity in size and susceptibility to deglycosylation seen in tryptase preparations. All three tryptase cDNAs are distinct from a recently reported cDNA obtained from a human lung mast cell library. A skin tryptase cDNA was used to isolate a human tryptase gene, the exons of which match one of the skin-derived cDNAs. The organization of the approximately 1.8-kilobase-pair tryptase gene is unique and is not closely related to that of any other mast cell or leukocyte serine protease. The 5' regulatory regions of the gene share features with those of other serine proteases, including mast cell chymase, but are unusual in being separated from the protein-coding sequence by an intron. High-stringency hybridization of a human genomic DNA blot with a fragment of the tryptase gene confirms the presence of multiple tryptase genes. These findings provide genetic evidence that human mast cell tryptases are the products of a multigene family.
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PMID:Human mast cell tryptase: multiple cDNAs and genes reveal a multigene serine protease family. 218 93

A second cDNA for human tryptase, called beta-tryptase, was cloned from a mast cell cDNA library in lambda ZAP. Its nucleotide sequence and corresponding amino acid sequence were determined and compared with those of a previously cloned tryptase cDNA, now called alpha-tryptase. The 1,142-base sequence of beta-tryptase encodes a 30-amino acid leader sequence of 3,089 D and a 245-amino acid catalytic region of 27,458 D. The amino acid sequence of beta-tryptase is 90% identical with that of alpha-tryptase, the first 20 amino acids of the catalytic portions being 100% identical. This identity, together with recognition of each recombinant protein by monoclonal antibodies directed against purified tryptase validate the tryptase identity of both alpha-tryptase and beta-tryptase cDNA molecules. Modest differences between the nucleic acid sequences of alpha- and beta-tryptase occurred throughout the cDNA molecules except in the 3' noncoding regions, which were identical. Although most highly conserved regions of amino acid sequence in the trypsin superfamily are conserved in both tryptase molecules, beta-tryptase has one carbohydrate binding site compared to two in alpha-tryptase, and one additional amino acid in the catalytic sequence. Regions of the substrate binding pocket in beta-tryptase (DSCQ, residues 218-221; SWG, residues 243-245) differ slightly from those in alpha-tryptase (DSCK, residues 217-220; SWD, residues 242-244). The presence of both alpha- and beta-tryptase sequences in each haploid genome was indicated by finding alpha- and beta-tryptase specific fragments after amplification by PCR of genomic DNA in 10 unrelated individuals. Localization of both alpha- and beta-tryptase sequences to human chromosome 16 was then performed by analysis of DNA preparations from 25 human/hamster somatic hybrids by PCR. It is now possible to assess the expression of each tryptase cDNA by mast cells and the relationship of each gene product to the active enzyme.
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PMID:Cloning and characterization of a second complementary DNA for human tryptase. 220 27


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