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)

[35S]Heparin was produced in vitro by incubation of rat peritoneal mast cells with [35S]sulfate and in vivo by injection of [35S]sulfate into rats. The [35S]heparin together with nonlabeled heparin in the mast cells was isolated in native form by mild methods that avoided the use of proteolytic enzymes or high alkali concentrations. The heparin had low anticoagulant activity. Incubations of mast cells with [35S]sulfate for less than several hours in vitro resulted in [35S]heparin of approximately Mr=200,000 to 400,000 based on gel filtration, while longer incubations yielded [35S]heparin of approximately Mr=750,000 that was similar to the nonlabeled heparin in the mast cells. When [3H]serine was included in the in vitro incubations, 3H-labeled material was found to co-chromatograph with the [35S]heparin. None of the heparin could be degraded by any of several proteolytic enzymes, but incubation for 14 h at 25 degrees with 0.5m NaOH degraded all samples to a size of approximately Mr=40,000. One-third of the [3H]serine label continued to co-chromatograph with the [35S]heparin after alkali treatment, while the remaining two-thirds appeared as smaller molecules completely separated from the [35S]heparin. Thus, native heparin of the mast cell may be an unusual proteoglycan that is resistant to proteolytic enzymes.
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PMID:Native heparin from rat peritoneal mast cells. 83 41

AMP deaminase (AMP aminohydrolase, EC 3.5.4.6) has been purified to apparent homogeneity from rat muscle. The preparation exhibits a single polypeptide band with a molecular weight of 60,000 on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The enzyme has a sedimentation coefficient of 11.3 S. Analysis by sedimentation equilibrium techniques showed the nat-ive enzyme to have a molecular weight of 238,000, whereas the enzyme, when analyzed in 6 M guanidine hydrochloride and 10 mM 2-mercaptoethanol, had a molecular weight of only 59,500. The amino acid composition of the enzyme was determined and peptide mapping was performed on a tryptic digest of S-carboxymethylated enzyme. NH2-terminal analysis by both the dansylation and cyanate procedures failed to identify a free NH2 terminus. Treatment of the enzyme with carboxypeptidase A resulted in the release of approximately 0.5 mol each of valine and leucine per 60,000 g of enzyme. The data presented indicate that hte native enzyme has a tetrameric structure consisting of four polypeptide chains each having a molecular weight of 60,000. The COOH-terminal analysis can be interpreted either as an indication of subunit heterogeneity or as a result of incomplete digestion of a -X-Leu-Val sequence at the end of a single type of polypeptide chain. Tryptic peptide maps strongly support the latter interpretation and suggest that the subunits are essentially identical.
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PMID:Rat muscle 5'-adenylic acid aminohydrolase. I. Purification and subunit structure. 115 74

In the course of experiments on the role of the COOH-terminal residues in pancreatic deoxyribonuclease, we undertook to ascertain whether the presence of sodium dodecyl sulfate would render the normally unavailable terminus susceptible to hydrolysis by carboxypeptidase A. When DNase A is dissolved in 0.005% sodium dodecyl sulfate the protein becomes enzymically inactive when assayed against DNA in the same sodium dodecyl sulfate concentration. The loss of activity caused by treatment with sodium dodecyl sulfate for 1 hour at 45 degrees can be fully restored if the detergent-containing solution is diluted 10-fold into 6 M guanidinium chloride and then 10-fold into a pH 7.0 buffer, 10 mM in CaCl2, prior to a 100-fold dilution for assay. The presence of Ca2+ is essential for the refolding process. If the same degree of dilution is made into sodium dodecyl sulfate-free buffer without the guanidinium chloride step, there is very little reversal of the inactivation. An almost complete loss of regenerable activity is caused by 1 hour of digestion by carboxypeptidase at 45 degrees in the presence of 0.03% sodium dodecyl sulfate. Although up to 6 amino acid residues can be removed from the COOH terminus, the loss of activity can be correlated with the removal of either 1 or 2 amino acid residues (-Leu-Thr) from the COOH-terminal sequence. Thus, DNase A is one of the several enzymes in which residues at the COOH terminus are essential to the active conformation. If the enzyme minus 2 to 6 terminal residues was mixed with a 15-residue COOH-terminal peptide (obtained by cyanogen bromide cleavage), only about 2% activity could be regenerated.
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PMID:Reversible inactivation of pancreatic deoxyribonuclease A by sodium dodecyl sulfate. Removal of COOH-terminal residues from the denatured protein by carboxypeptidase A. 116 40

Inhibitors of the peptidase and esterase activities of carboxypeptidases A and B have been isolated from extracts of Ascaris lumbricoides var suis. These proteins were obtained by treatment of the aqueous extracts at low pH, precipitation with ammonium sulfate, molecular sieving on Bio-Gel P-4, and chromatography on DEAE-cellulose. The inhibitors were resolved into three homogeneous peaks on CM-cellulose. These components, CM-A, CM-B, and CM-C, have constant specific activity and were recovered in a 41% yield. They moved as single bands when subjected to electrophoresis at high or low pH on polyacrylamide gels and they have similar amino acid compositions. Methionine, tyrosine, and cysteine are absent from each of the inhibitors. The 65 residues of CM-B suggest a minimum molecular weight of 7530, in close agreement to the value of 7600 +/- 200 determined on a Bio-Gel P-100 column. Each of the proteins has the same NH2-terminal residues, NH2-Asx-Glx-Val-Glx- and the same COOH-terminal residue, leucine. A plot of per cent acrylamide versus log relative mobility suggests that the three proteins are charge isomers. CM-B appears to be stable to high NaCl concentrations, extremes of pH, high temperatures, and digestion by intestinal proteases. Carboxypeptidase C, carboxypeptidase N, and yeast protease C are not inhibited by CM-B. However, the exopeptidase and esterase activities of human carboxypeptidase A are inhibited. The inhibitors appear to bind to bovine carboxypeptidase A with an atypical stoichiometry. Two moles of CM-B inhibitor bind to 1 mol of enzyme. The evidence is: (a) a demonstrated purity of bovine carboxypeptidase A, (b) minimal and maximal inhibitor molecular weights by different methods, of 7600 and 8300, and (c) a maximum specific activity of apparently homogeneous inhibitors which is 50% of that predicted for unit stoichiometry.
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PMID:Characterization of proteins from Ascaris lumbricoides which bind specifically to carboxypeptidase. 126 22

An animal model for malignant mastocytosis is described in mice reconstituted with bone marrow cells expressing the v-erbB oncogene. The lethal mast cell disease is characterized by massive infiltration of bone marrow, spleen, and several other visceral organs by connective tissue mast cells, which normally reside in the skin and the peritoneal cavity. As is frequently found in malignant mastocytosis, the v-erbB-induced mast cell disease was accompanied in some primary recipients by an acute myelogenous leukemia (AML) that killed all secondary recipients regardless of whether the AML was already evident in the primary host. The infiltrating mast cells stained strongly positive with berberine sulfate, suggesting that they were terminally differentiated and in vitro they showed only a weak proliferative capacity. The leukemias were clonal but apparently of different origin than the malignant mast cells, implying the transformation of two independent cell populations. Leukemic cells expressed various myeloid-specific markers as well as the B220 antigen, normally associated with the B-cell lineage. However, the Ig heavy chain genes were still in germ line configuration. In culture, these cells proliferated in the absence of exogenous growth factors and had the capacity to differentiate into mature myeloid cells. Preliminary experiments suggest that v-erbB may use parts of a signal transduction pathway normally coupled to the c-kit receptor. The v-erbB-induced malignant mast cell disease should provide a useful animal model for elucidating the cause for malignant mastocytosis in humans and to explore possible therapeutic strategies.
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PMID:Development of a lethal mast cell disease in mice reconstituted with bone marrow cells expressing the v-erbB oncogene. 135 Jul 40

Hemopoietic stem cell factor (SCF), which is the ligand for the proto-oncogene c-kit receptor (allelic with W locus) and the product of Sl locus of the mouse, has recently been cloned. The human homologue has also been cloned, and recombinant protein (human rSCF) expressed and purified to homogeneity. To determine the effect of human rSCF in the presence or absence of human rIL-3 on human bone marrow-derived mast cells and basophils, human CD34+ pluripotent progenitor cells, highly enriched (greater than 99%) from bone marrow mononuclear cells, were cultured over agarose surfaces (interphase cultures) in the presence of human rIL-3, human rIL-3 and increasing concentrations of human rSCF, or human rSCF alone. Over 3 to 4 wk, human rSCF acted synergistically with human rIL-3 at all concentrations, producing a three- to fivefold increase in total, mast cell, and basophil numbers over human rIL-3 alone when used at 100 ng/ml. The percentage of cell types in the human rIL-3 and human rIL-3 plus human rSCF cultures, however, remained the same, with basophils constituting 18 to 35% of the final cultured cells, and mast cells 3% or less of the final cell number. In the presence of human rSCF alone, the combined total percentage of mast cells and basophils was 0 to 1.0%, the majority of cells being macrophages. Mast cells cultured in human rIL-3 plus human rSCF, but not human rIL-3 alone, were berberine sulfate positive, suggesting the presence of heparin proteoglycans within granules. Electron microscopic examination of cultures supplemented with human rIL-3 and rSCF, but not human rIL-3 alone, revealed that after 3 wk in culture, mast cell granules contained tryptase and exhibited scroll, reticular, and homogeneous patterns as seen previously in CD34+/3T3 fibroblast cocultures. Thus, CD34+ cells cultured in the presence of both human rIL-3 and rSCF give rise to cultures containing increased numbers of basophils and mast cells, with the mast cells by ultrastructural studies showing evidence of maturation although the percentages of basophils and mast cells arising in these cultures remained unchanged.
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PMID:Effect of IL-3 and stem cell factor on the appearance of human basophils and mast cells from CD34+ pluripotent progenitor cells. 137 May 17

The proliferative capacity of mouse connective tissue-type mast cells (CTMC) was analyzed by using a newly discovered c-kit ligand, termed stem cell factor (SCF). More than 90% of CTMC in the peritoneal cavity responded to recombinant rat SCF (rrSCF) and were able to give rise to pure mast cell colonies in methylcellulose culture. Serial observation (mapping) of growth of individual CTMC in culture containing rrSCF confirmed their striking proliferative ability. No serum but accessory cells (non-CTMC cells) in the peritoneal population were required for the clonal growth of CTMC induced by rrSCF in our methylcellulose culture of whole peritoneal cells. The rrSCF-induced mast cell colony formation from peritoneal CTMC was completely inhibited by the addition of anti-c-kit antibody, which can block the binding of SCF to c-kit, to the culture. When IL-3 was combined with rrSCF, mast cell colonies dramatically increased in size. Mapping studies revealed that the combination of the two factors augmented the proliferative rate of CTMC. Approximately 60% of the constituent cells of the mast cell colonies which were formed from peritoneal CTMC in the culture containing rrSCF alone were stained with berberine sulfate, which is a characteristic of CTMC. However, most mast cells which were induced by rrSCF+IL-3 from peritoneal CTMC contained berberine(-)-safranin(-)-Alcian blue(+) granules. Although IL-4 exhibited little synergism with rrSCF in the induction of CTMC proliferation, the addition of IL-4 to the culture containing rrSCF+IL-3 resulted in an increase in mast cells which retained CTMC characteristics.
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PMID:Stimulation of mouse connective tissue-type mast cells by hemopoietic stem cell factor, a ligand for the c-kit receptor. 137 44

Immunization of BALB/c mice with denatured DNA (dnDNA)-methylated bovine serum albumin (MBSA) complex along with aluminium hydroxide gel as adjuvant, resulted in the induction of anti-DNA antibodies of both IgG and IgE isotypes demonstrable by avidin-biotin micro enzyme-linked immunosorbent assay (ELISA) and solid phase radioimmunoassay (SPRIA), respectively. In contrast to the high levels of IgG2a and IgG2b anti-DNA antibodies observed in SLE-prone autoimmune mice, more than 90% of the anti-DNA antibodies of IgG isotype were found to be of IgG1 subclass. Specificity of both IgG and IgE antibodies which recognized activated DNA, dnDNA and double-stranded DNA but not RNA was established by competitive ELISA and SPRIA inhibition assays. These antibodies cross-reacted with cibacron blue and chondroitin sulfate but not with various other proteoglycans, nucleosides and nucleotides. Passive cutaneous anaphylaxis reaction in rats showed that these antibodies are capable of inducing in vivo degranulation of mast cells in a dose-dependent manner. These studies lend support to the concept that IgE antibodies directed against DNA may mediate mast cell degranulation and thus contribute to immediate-type hypersensitivity phenomena including hives seen in patients with systemic lupus erythematosus and to the localization of IgE-nucleic acid complexes.
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PMID:Induction of anti-DNA IgG and IgE antibodies in BALB/c mice. 141 1

Chymase, a potent secretagogue for airway gland serous cells, is stored in secretory granules and released from mast cells together with proteoglycans. To investigate the hypothesis tha tproteoglycans modulate chymase-induced effects, we studied the influence of proteoglycans purified from dog mastocytoma cells on chymase-induced secretion from cultured bovine airway gland serous cells. Heparin proteoglycans reduced the chymase-induced secretory response, whereas glycosaminoglycans and chondroitin sulfate proteoglycans had less of an effect. Chymase released together with proteoglycans from activated mast cells caused secretion comparable to that caused by purified chymase reconstituted with purified proteoglycans. Despite partial inhibition by exocytosed proteoglycans, the secretagogue activity of chymase remains substantial compared to that of histamine. However, proteoglycans virtually abolished chymase-induced degradation of the products of serous cell secretion. Although the secretagogue and proteoglycanase activities of chymase are inhibited by most classes of mast cell granule-associated glycans, the amidolytic activity of chymase toward tripeptide 4-nitroanilide substrates is augmented. These findings suggest that mast cell proteoglycans modulate the secretagogue, proteoglycanase, and peptidase activity of chymase, and the results predict that the extent of this modulation in vivo depends on the nature of the proteoglycans with which chymase is released from mast cells.
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PMID:Mast cell proteoglycans modulate the secretagogue, proteoglycanase, and amidolytic activities of dog mast cell chymase. 157 74

The knowledge about the differentiation of basophilic leukocytes is fragmentary. This report discusses a detailed phenotypic characterization of molecular markers for hematopoietic differentiation in a basophilic leukemia cell line, KU812. The expression of markers for lymphoid, erythroid, neutrophil, eosinophil, monocytic, megakaryocytic, mast cell and basophil differentiation was analyzed at the mRNA level by Northern blots in the KU812 cells, and for reference, in a panel of human cell lines representative of the different hematopoietic differentiation lineages. KU812 was found to express a number of mast cell and basophil-related proteins, i.e. mast cell tryptase, mast cell carboxypeptidase A, high-affinity immunoglobulin (IgE) receptor alpha and gamma chains and the core protein for heparin and chondroitin sulphate synthesis. We found no expression of a number of monocyte/-macrophage or neutrophil leukocyte markers except for lysozyme. From earlier studies, it has been shown that lysozyme is not expressed in murine mucosal mast cell lines. This finding, together with the expression of the mast cell carboxypeptidase in KU812 might distinguish the phenotype of this cell line from that typical of mucosal mast cell lines in rodents. We found a low level of expression of the eosinophil and basophil marker, major basic protein, which might indicate a relationship between basophils and eosinophils. No expression is, however, detected with the eosinophil-specific markers eosinophil cationic protein, eosinophil-derived neurotoxin or eosinophil peroxidase. We also report an extensive screening for inducers of basophilic differentiation of the KU812 cells. The most efficient protocol of induction included serum starvation which led to a dramatic increase in a number of markers specific for mast cells and basophils such as tryptase, carboxypeptidase A and the heparin core protein. Finally, diisopropylfluorophosphate analysis of total protein extracts from KU812 show four labeled protein bands with sodium dodecyl sulfate-polyacrylamide gel electrophoresis, indicating that this cell line expresses at least three previously undescribed serine proteases of which one or more could be a potential basophil-specific marker(s).
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PMID:Phenotypic characterization of KU812, a cell line identified as an immature human basophilic leukocyte. 163 3


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