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)

The neuronal membrane protein which binds the K+-channel ligands dendrotoxin, mast cell degranulating peptide, and beta-bungarotoxin was purified from rat brain membranes. When analysed on 10% SDS gel electrophoresis, the purified protein contained two peptides: the toxin-binding subunit of apparent Mr 90,000 and another peptide of Mr 38,000. Neuraminidase treatment reduced the Mr of the toxin-binding subunit to 70,000. Glycopeptidase F gave a further reduction to Mr 65,000. In contrast, the peptide of Mr 38,000 showed no change in Mr upon treatment with neuraminidase and/or glycopeptidase F. It is concluded that the toxin-binding subunit of the dendrotoxin-binding protein, a presumptive K+ channel, is a sialated membrane protein with a peptide core of, at most, Mr 65,000.
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PMID:Enzymatic deglycosylation of the dendrotoxin-binding protein. 270 49

The azurophil granules of neutrophil granulocytes contain neutral proteases such as leukocyte elastase and cathepsin G. These are synthesized as inactive precursors, but following proteolytic processing, they are stored in granules as active enzymes. We describe the establishment of a transgenic cellular model for expression of the human myeloid serine protease cathepsin G. The cDNA for preprocathepsin G was stably expressed in the rat basophilic/mast cell line RBL-1 and the translation product was characterized by use of biosynthetic labeling followed by immunoprecipitation, SDS-polyacrylamide gel electrophoresis, and fluorography. Conversion into complex form of an asparagine-linked carbohydrate unit of approximately 3.5 kDa was shown, as judged by the products obtained upon treatment with endoglycosidase H and N-glycanase. Proteolytic processing of 32.5-kDa procathepsin G into a 31-kDa form, within 1-2 h after synthesis, was demonstrated by pulse-chase experiments. Further processing into a 30-kDa form also occurred to a minor extent. The processed forms were enzymatically active, as judged by affinity for the serine protease inhibitors diisopropylfluorophosphate and aprotinin. Translocation of processed forms of cathepsin G to high density fractions, indicating targeting of the protease to granules, was demonstrated by subcellular fractionation. The weak base NH4Cl was shown to delay the processing and enzymatic activation of cathepsin G, whereas the monovalent ionophore monensin completely inhibited both events. Our data demonstrate that human cathepsin G transfected to rat RBL-1 cells, is proteolytically processed into enzymatically active forms and that subcellular transfer to granular organelles occurs. As the processing of transgenic human cathepsin G corresponds to that of endogenous protease of myeloid cells, the model should provide new unique possibilities to further characterize the activation and granular targeting of myeloid serine proteases.
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PMID:Processing of human cathepsin G after transfection to the rat basophilic/mast cell tumor line RBL. 792 11

Although the steady-state level of the mouse mast cell protease (mMCP) 7 transcript is below detection in the serosal and mucosal mast cells of the BALB/cJ mouse, the IL-3-dependent, bone marrow-derived mast cells (mBMMC) of this strain and four other strains contain a high steady-state level of the mMCP-7 transcript. To further analyze the expression of this mast cell tryptase, a mMCP-7-specific IgG was obtained by immunizing a rabbit with a 19-residue synthetic peptide that corresponds to its unique amino acid sequence at residues 160 to 178 (anti-mMCP-7(160-178). In a SDS-PAGE/immunoblot analysis of lysates of BALB/cJ mBMMC, anti-mMCP-7(160-178) IgG recognized a diffuse 31- to 36-kDa protein, which shifted to a sharp 27-kDa protein after treatment with N-glycanase. As assessed immunohistochemically, mMCP-7 protein is present not only in the secretory granules of BALB/cJ mBMMC, but also in the ear mast cells of this strain. In contrast, the ear mast cells of the C57BL/6J mouse do not contain detectable levels of mMCP-7 protein, although the ear mast cells of both mouse strains contain mMCP-5 protein. Because mMCP-7 mRNA and protein also were not detected in mBMMC from the C57BL/6J mouse, the failure of the ear mast cells of this strain to express mMCP-7 is most likely a consequence of an intrinsic abnormality in the mast cell-committed progenitor cells themselves, or in the bone marrow microenvironment that induces its mast cell progenitor cells to express this tryptase.
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PMID:Lack of expression of the tryptase mouse mast cell protease 7 in mast cells of the C57BL/6J mouse. 807 72

There has long been conjecture over the degree to which there may be structural and functional heterogeneity in the tetrameric serine protease tryptase (EC 3.4.21.59), a major mediator of allergic inflammation. We have applied 2D gel electrophoresis to analyze the extent, nature, and variability of this heterogeneity in lysates of mast cells isolated from lung and skin, and in preparations of purified tryptase. Gels were silver stained, or the proteins transferred to nitrocellulose blots and probed with either tryptase-specific monoclonal antibodies or various lectins. Tryptase was the major protein constituent in mast cell lysates, and presented as an array of 9-12 diffuse immunoreactive spots with molecular masses ranging from 29 to 40 kDa, and pI values from 5.1 to 6.3. Although the patterns obtained for lung and skin tryptase were broadly similar, differences were observed between tissues and between individual donors. Lectin binding studies indicated the presence of mono-antennary or bi-antennary complex-type oligosaccharide with varying degrees of sialylation. Deglycosylation with protein-N-glycosidase F (PNGase F) reduced the size of both lung and skin tryptase, while incubation with PNGase F or neuraminidase narrowed the pI range, indicating variable degrees of glycosylation as a major contributor to the size and charge heterogeneity. Comparison of different purified preparations of lung and skin tryptase revealed no significant difference in pH profiles, but differences were seen in reactivity towards a range of chromogenic substrates, with substantial differences in Km, kcat and degree of cooperativity. Mathematical modeling indicated that the variety in kinetics parameters could not result solely from the sum of varying amounts of isoforms obeying Michaelis-Menten kinetics but with different values of Km and kcat. The heterogeneity demonstrated for tryptase in these studies suggests that there are important differences in tryptase function in different tissues.
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PMID:The heterogeneity of mast cell tryptase from human lung and skin. 1260 78

Extracellular and cell surface proteins are generally modified with N-linked glycans and glycopeptide enrichment is an attractive tool to analyze these proteins. The role of N-linked glycoproteins in cardiovascular disease, particularly ischemia and reperfusion injury, is poorly understood. Observation of glycopeptides by mass spectrometry is challenging due to the presence of abundant, nonglycosylated analytes, and robust methods for purification are essential. We employed digestion with multiple proteases to increase glycoproteome coverage coupled with parallel glycopeptide enrichments using hydrazide capture, titanium dioxide, and hydrophilic interaction liquid chromatography with and without an ion-pairing agent. Glycosylated peptides were treated with PNGase F and analyzed by liquid chromatography-MS/MS. This allowed the identification of 1556 nonredundant N-linked glycosylation sites, representing 972 protein groups from ex vivo rat left ventricular myocardium. False positive "glycosylations" were observed on 44 peptides containing a deamidated Asn-Asp in the N-linked sequon by analysis of samples without PNGase F treatment. We used quantitation via isobaric tags for relative and absolute quantitation (iTRAQ) and validation with dimethyl labeling to analyze changes in glycoproteins from tissue following prolonged ischemia and reperfusion (40 mins ischemia and 20 mins reperfusion) indicative of myocardial infarction. The iTRAQ approach revealed 80 of 437 glycopeptides with altered abundance, while dimethyl labeling confirmed 46 of these and revealed an additional 62 significant changes. These were mainly from predicted extracellular matrix and basement membrane proteins that are implicated in cardiac remodeling. Analysis of N-glycans released from myocardial proteins suggest that the observed changes were not due to significant alterations in N-glycan structures. Altered proteins included the collagen-laminin-integrin complexes and collagen assembly enzymes, cadherins, mast cell proteases, proliferation-associated secreted protein acidic and rich in cysteine, and microfibril-associated proteins. The data suggest that cardiac remodeling is initiated earlier during reperfusion than previously hypothesized.
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PMID:Quantitative N-linked glycoproteomics of myocardial ischemia and reperfusion injury reveals early remodeling in the extracellular environment. 2144 15