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)

Exogenous addition of purified chymase, a rat serosal mast cell (RSMC) chymotryptic enzyme, results in RSMC degranulation at 37 degrees, but not at 1 degree. Chymase can cause an active site-dependent inducing event at 1 degree such that RSMC degranulation occurs if the cells are later incubated at 37 degrees. RSMC exposed to chymase or other stimuli were surface radiolabelled using 125I and Iodo-Gen, solubilized with 1% Nonidet-40, and the resulting 25,000 g supernatants analysed by SDS-PAGE and autoradiography. A 125I-labelled RSMC membrane protein of approximate 90,000 MW decreased upon exposure to either chymase or alpha-chymotrypsin (alpha-CT) for 5 min at 37 degrees or to chymase for 60 min at 1 degree. Exposure of RSMC to the secretagogues ionophore A23187, compound 48/80, and anti-IgE for 5 min at 37 degrees resulted in beta-hexosaminidase (a secretory granule enzyme) release, but did not cause a detectable change in the 90,000 MW surface-labelled protein. Lima bean trypsin inhibitor, which inhibits both the esterase and RSMC degranulation activities of chymase and alpha-CT, prevented the disappearance of the 125I-labelled 90,000 MW band when added with chymase or alpha-CT. Exposure of RSMC to chymase at 1 degree for 0-10 min, prior to addition of LBTI, led to a progressive disappearance of the 90,000 MW band, which corresponded to the kinetics of priming for subsequent RSMC degranulation at 37 degrees. When RSMC were exposed to trypsin (2.5 micrograms/ml) for 0-120 min at 1 degree, a progressive disappearance of the 90,000 MW band occurred, in association with a loss of sensitivity to subsequent activation by chymase at 37 degrees. The disappearance of the 90,000 MW determinant in association with chymase-mediated priming for degranulation and the inability of chymase to mediate degranulation of trypsin-treated RSMC, which lack this membrane protein, suggests that it is involved in chymase-mediated RSMC degranulation.
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PMID:Cleavage of a rat serosal mast cell membrane component during degranulation mediated by chymase, a secretory granule protease. 231 65

The structure of serine carboxypeptidase II from wheat bran has been determined to 3.5-A resolution by multiple isomorphous replacement, solvent flattening, and crystallographic refinement. The amino acid sequence has been fit to the electron density map and the model refined to a conventional crystallographic R factor of 20.9%. The molecule is an alpha + beta protein and contains a "catalytic triad" (Asp338, His397, and Ser146) similar in arrangement to those in chymotrypsin and subtilisin. The -fold of the polypeptide backbone is, however, completely different from those enzymes. This suggests that this is a third example of convergent evolution to a common enzymatic mechanism. The -fold is, on the other hand, surprisingly similar to that of the zinc proteinase carboxypeptidase A.
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PMID:Structure of wheat serine carboxypeptidase II at 3.5-A resolution. A new class of serine proteinase. 232 88

In inside-out red cell membrane vesicles active calcium transport and the formation of the enzyme-phosphate complex (EP) of the calcium pump were simultaneously investigated and the effects of a limited proteolytic digestion examined. In order to visualize the proteolyzed EP forms we have induced the formation of a maximum level EP from [gamma-32P]ATP in the presence of Ca2+ + La3+ and applied a good-resolution acidic discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoresis system. Proteolysis of inside-out vesicle membranes by trypsin, Pronase, papain, or chymotrypsin produces a calmodulin-like activation of the calcium pump, abolishes its calmodulin sensitivity, and decreases the original 140-kDa EP complex to a limit polypeptide of 80 kDa. Trypsin digestion produces another major intermediary fragment of 90 kDa, which is still a low-activity calmodulin-sensitive form of the pump. The red cell calcium pump is activated by trypsin both in the absence and presence of Ca2+ during digestion although the rate of activation and the appearance of the 80-kDa polypeptide are enhanced by Ca2+. If proteolytic digestion is carried out by chymotrypsin, a calmodulin-insensitive maximum activation of the calcium pump coincides with the formation of a 125-130-kDa EP-forming polypeptide. Chymotrypsin and carboxypeptidase A have synergistic effects on the formation of this latter high-activity species. Based on these data we suggest a probable molecular arrangement for the functional parts of the red cell membrane calcium pump.
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PMID:Molecular characterization of the in situ red cell membrane calcium pump by limited proteolysis. 242 14

The appearance and activity of various porcine pancreatic hydrolases were studied during fetal and postnatal development. Quantitatively, the enzyme activities in activated pancreas homogenates were low but increased during the second half of the fetal period, using the substrates Bz-Arg-pNA for measuring anodal and cathodal trypsin, Suc-Phe-pNA (chymotrypsin A and C, and elastase II) and Suc-(Ala)3-pNA (elastase I and protease E). Postnatally, after an initial decrease during the first week, the enzyme activities increased markedly, especially from 10-14 weeks to 6 months. The individual hydrolases were identified after electrophoretic separation in agarose gel and staining with various substrates either directly in the gel or after transfer to nitrocellulose membranes (enzymoblotting). During the fetal period, chymotrypsin A and B, elastase II, carboxypeptidase A, and amylase appeared at approximately 65 days and anodal trypsin, at approximately 76 days. After birth, new proteinases appeared after the first week including chymotrypsin C, cathodal trypsin, and protease E, whereas elastase I was found from 5 weeks after birth. Concomitantly, unidentified "fetal proteinase(s)" with caseinolytic, Ac-Phe-beta NE and CBZ-Ala-beta NE activities began to diminish and disappeared 10-14 weeks after birth. This study showed a marked increase in the overall pancreatic enzyme activities, as well as an age-dependent expression of the variety of pancreatic hydrolases during porcine ontogeny.
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PMID:Development of porcine pancreatic hydrolases and their isoenzymes from the fetal period to adulthood. 244 72

The relationship between inter-alpha inhibitor (I alpha I) and urinary proteinase inhibitor (UPI) was examined by comparing purified UPI with a proteolytic fragment of I alpha I (I'), and by demonstrating that inflammatory cells produce similar fragments under physiologic conditions. Purified I', derived by chymotrypsin digestion of I alpha I, was similar to UPI in apparent molecular weight (68,000-69,000), amino acid composition, immunoreactivity, and inhibitory activity against trypsin, chymotrypsin, and neutrophil elastase. The production of similar inhibitory fragments by murine peritoneal macrophages, human neutrophils, and a murine mast cell line was quantified. Neutrophils were most efficient at proteolyzing I alpha I. Comparison of the pattern of I alpha I degradation by neutrophil preparations with that by pure enzymes, suggested that both elastase and cathepsin G mediate neutrophil proteolysis of I alpha I. These proteinases may thus be responsible for inflammation-related increases in UPI-like inhibitor levels in vivo.
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PMID:Inflammatory cells degrade inter-alpha inhibitor to liberate urinary proteinase inhibitors. 246 21

A fluorescent peptide substrate to explore the protease specificity for the amino acid regions C- and N-terminal to the cleavage site has been designed. Intramolecular quenching of indole fluorescence by an N-terminal dansyl group separated by six amino acid residues forms the basis of this assay. For a particular enzyme, specificity can be designed into the peptide sequence by means of the number of residues that separate the two chromophores. In the present instance, the heptapeptide Dns-Gly-Lys-Tyr-Ala-Pro-Trp-Val is used to assay angiotensin converting enzyme (ACE), Astacus protease, carboxypeptidase A, alpha-chymotrypsin, and trypsin, all of which cleave the peptide in accord with their known specificity: Trypsin and Astacus protease hydrolyze only the Lys-Tyr and Tyr-Ala bonds, respectively. alpha-Chymotrypsin primarily cleaves the Tyr-Ala bond while ACE makes three successive dipeptidyl cleavages from the C-terminus. Carboxypeptidase rapidly hydrolyzes first the Trp-Val and then the Pro-Trp bond. For all of the enzymes, catalytic activity (kcat/Km) is in the range from 10(5) to 10(6) M-1 s-1. Hydrolysis causes a fluorescence increase in the 310 to 410 nm region of 8.6- to 13.6-fold depending on the enzyme that is assayed. Assays can be designed based on the increase in tryptophan fluorescence or by individual product analyses using thin-layer or high-performance liquid chromatography. The specificity and sensitivity of such internally quenched fluorescent oligopeptides would seem to be ideal for the assay of specific endoproteases.
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PMID:A fluorescent oligopeptide energy transfer assay with broad applications for neutral proteases. 255 28

Hydrolysis of ursodeoxycholyl-p-aminobenzoic acid (PABA-UDCA), a synthetic bile acid conjugate used for the evaluation of the activity of intestinal bacterial growth, was studied with pancreatic enzymes, carboxypeptidase A, carboxypeptidase B, trypsin alpha-chymotrypsin, cholylglycine hydrolase, liver homogenate, small intestinal homogenates, and plasma, in comparison with the hydrolysis of glycocholic acid, ursodeoxycholyl-L-leucine (L-Leu-UDCA), and ursodeoxycholyl-L-lysine (L-Lys-UDCA). PABA-UDCA was specifically cleaved by bacterial cholylglycine hydrolase to ursodeoxycholic acid and para-aminobenzoic acid (PABA), but not by pancreatic enzymes. L-Leu-UDCA was cleaved by pancreatic enzymes, carboxypeptidase A, and cholylglycine hydrolase. L-Lys-UDCA was cleaved by pancreatic enzymes, carboxypeptidase B, and cholylglycine hydrolase. The small amount of glycocholic acid was cleaved by pancreatic enzymes and carboxypeptidase A and B, and cholylglycine hydrolase hydrolyzed glycocholic acid completely. In everted gut sac experiments, PABA-UDCA was absorbed by active transport in the rat terminal ileum, and the same rate of PABA was absorbed by passive diffusion in the four segments of the rat small intestine. These observations indicate that PABA-UDCA test can evaluate the activity of small intestinal bacterial growth.
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PMID:Hydrolysis and absorption of a conjugate of ursodeoxycholic acid with para-aminobenzoic acid. 263 32

The carboxypeptidase A inhibitor from Ascaris suum was isolated from aqueous extracts by affinity chromatography toward immobilized carboxypeptidase A. The amino acid sequence is DQVRKCLSDT10DCTNGEKCVQ20KNKICSTIVE30IQRCEKEHFT40IPCKSNNDCQ50VWAHEKICN K60LPWGL65 . The carboxypeptidase A inhibitor is not homologous with the chymotrypsin/elastase or trypsin inhibitors from Ascaris, but shows homology in a 9-residue internal sequence with the 37/39-residue carboxypeptidase inhibitors from tomato and potato. The carboxy-terminal 5 (4) residues in the three inhibitors are similar, suggesting a common mechanism of inhibition.
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PMID:Carboxypeptidase inhibitors from Ascaris suum: the primary structure. 264 95

The amino acid sequence has been determined of a mouse mucosal mast cell protease isolated from the small intestines of mice infected with Trichinella spiralis. The active protease contains 226 residues. Those corresponding to the catalytic triad of the active site of mammalian serine proteases (His-57, Asp-102, and Ser-195 in chymotrypsin) occur in identical positions. A computer search for homology indicates 74.3% and 74.1% sequence identity of the mouse mast cell protease compared to those of rat mast cell proteases I and II (RMCP I and II), respectively. The six half-cystine residues in the mouse mast cell protease are located in the same positions as in the rat mast cell proteases, cathepsin G, and the lymphocyte proteases, suggesting that they all have identical disulfide bond arrangements. At physiological pH, the mouse and rat mucosal mast cell proteases have net charges of +3 and +4, respectively, as compared to +18 for the protease (RMCP I) from rat connective tissue mast cells. This observation is consistent with the difference in solubility between the mucosal and connective tissue mast cell proteases when the enzymes are extracted from their granules under physiological conditions.
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PMID:Amino acid sequence of a mouse mucosal mast cell protease. 270 64

Biochemical information regarding the mechanism of amide bond hydrolysis offers insight into the possible chemical groups in the enzyme active site responsible for hydrolysis. Assuming that these groups have a relatively fixed geometry in accord with their functional role, then their three-dimensional position in space can be determined if sufficient structural diversity exists within the data set of compounds with known affinities. Each compound which binds can be augmented by additional chemical groups to represent the receptor's functional groups. For each compound, the set of geometrical arrangements of these groups which would show optimal binding to the compound can be determined by systematic search. A common geometric arrangement representing the active site geometry should be present for each compound. In studies of the binding of mechanism-based inhibitors of chymotrypsin, Naruto et al. (1985) showed some movement of active site residues to accommodate different ligands. Nevertheless, this procedure found a unique active site geometry for ACE which compared favorably with that of carboxypeptidase A, an enzyme of analogous function.
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PMID:Mechanism-based analysis of enzyme inhibitors of amide bond hydrolysis. 272 59


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