Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Peptide boronic acids, such as methoxysuccinyl-Ala-Ala-Pro-(L)boro-Phe-OH, its pinacol ester, and t-butyloxycarbonyl-Phe-Pro-(L)boro-Phe-pinacol, inhibited the activity of chymase from connective tissue mast cells approximately 40- to 80-fold more than atypical chymase from mucosal mast cells, and did not inhibit trypsin. Only peptide boronic acids containing "L" forms of boronic acids were inhibitory. The Ki values of these peptide boronic acids for chymase were in the 60-170 nM concentration range, like those of the natural inhibitors tested, but all the natural inhibitors tested except Eglin C and chymostatin inhibited both chymase and trypsin. Thus these peptide boronic acids should be useful for selective inhibition of chymase with less inhibitory activity for atypical chymase and without inhibition of trypsin. These peptide boronic acids markedly inhibited histamine release induced by anti-rat immunoglobulin E, suggesting that chymase in connective tissue mast cells plays some role in the process of histamine release. These peptides are assumed to be therapeutically useful for treatment of allergic inflammations catalyzed by chymase.
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PMID:Peptide boronic acids, substrate analogs, inhibit chymase, and histamine release from rat mast cells. 246 Apr 36

Serine proteases in mast cell granules, such as chymase, atypical chymase, and tryptase, which are major proteins in the granules, may play important roles in the process of immunoglobulin E (IgE)-mediated degranulation and in pathobiological alterations in tissues. Indeed, inhibitors of chymase, substrate analogs, and antichymase F(ab')2, but not inhibitors of tryptase, markedly inhibited histamine release induced by IgE-receptor bridging but not that induced by Ca ionophore. In contrast, inhibitors of metalloprotease inhibited histamine release induced not only by IgE-receptor bridging but also by Ca ionophore. These results suggest that chymase and metalloprotease are involved at different steps in the process of degranulation. The extents of inhibition of histamine release were closely correlated with the amounts of the inhibitors of chymase accumulated in the granules. After degranulation, the released proteases may in part contribute to pathobiological alterations in allergic disorders through generations of C3a anaphylatoxin and thrombin by human and rat tryptase, respectively, and those of angiotensin II and a chemotactic factor of neutrophils by human and rat chymase, respectively. Moreover, chymase and atypical chymase from rat were shown to destroy type IV collagen, and human tryptase was found to hydrolyze various plasma proteins, such as fibrinogen and high-molecular-weight kininogen. The biological activities of tryptase and chymase from rat may be regulated by their dissociation from and association with trypstatin, an endogenous inhibitor of these proteases.
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PMID:Biological functions of serine proteases in mast cells in allergic inflammation. 246 15

Human pulmonary mast cells contain the serine proteases tryptase and chymase. Chymase is present in much smaller quantities than tryptase. The definite physiological role of both enzymes remains to be elucidated, angiotensin processing has been proposed as one possible function of chymase. A dose-dependent inhibition of A 23187-induced histamine release from dispersed human lung mast cells was observed after pretreatment with diisopropylfluorophosphate (DFP) or 1-1-tosyamide-2-phenylethyl chloromethyl ketone (TPCK) but not with N-2-p-tosyl-1-lysine chloromethyl ketone (TLCK). In contrast, no inhibition was observed under the same conditions with isolated rat peritoneal mast cells. These results indicate that a chymase is probably an important factor in a late phase of human lung mast cell activation. Current work focuses on the isolation of human lung chymase to further investigate this topic.
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PMID:The role of chymase in ionophore-induced histamine release from human pulmonary mast cells. 246 1

Submucosal glands are the major sources of airway secretions in most mammals. Mast cells are abundant in the environment of airway submucosal glands and are rich sources of secreted proteases. To investigate the hypothesis that mast cell proteases stimulate airway gland secretion, we studied the ability of the two major mast cell granule proteases, chymase and tryptase, to cause secretion of 35S-labeled macromolecules from a line of cultured bovine airway gland serous cells. Mast cell chymase and tryptase were purified from dog mastocytoma cells. Chymase markedly stimulated serous cell secretion in a concentration-dependent fashion with a threshold of 10(-10) M, whereas tryptase had no effect. The response to 10(-8) M chymase (1530 +/- 80% over base line) was approximately 10-fold higher than that evoked by other agonists such as histamine and isoproterenol. The predominant 35S-labeled macromolecule released by chymase was chondroitin sulfate proteoglycan, the glycoconjugate present in serous cell secretory granules. The response to chymase was non-cytotoxic and was blocked by active site inhibitors of chymase (soybean trypsin inhibitor and chymostatin) and by inhibitors of cellular energy metabolism (azide,2,4-dinitrophenol, dicumarol). Supernatant obtained by degranulation of mastocytoma cells caused a secretory response of comparable magnitude to that caused by chymase. These findings demonstrate that chymase, but not tryptase, is a potent secretagogue for airway gland serous cells, and they suggest a possible role for chymase-containing mast cells in the pathogenesis of airway hypersecretion.
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PMID:Mast cell chymase. A potent secretagogue for airway gland serous cells. 249 59

Recent evidence suggests that nonadrenergic airway relaxation may be controlled by vasoactive intestinal peptide (VIP). The magnitude and duration of smooth muscle relaxation in response to VIP may be influenced by rates of peptide degradation after release from efferent peptidergic neurons. To explore the potential role of mast cell mediators in modulating neural control of airway tone, we studied the effect of the mast cell proteases tryptase and chymase on airway smooth muscle relaxation induced by VIP in ferret airway. Tracheal rings precontracted by serotonin (10(-6) M) in a muscle bath were relaxed by VIP (10(-7) M). We found that protease-rich supernatant obtained by degranulation of dog mastocytoma cells reversed VIP-induced relaxation, as did highly purified tryptase and chymase incubated with the tracheal rings. Either enzyme completely reversed the effect of VIP, but tryptase was more potent than chymase, paralleling previous test tube observations on the relative rates of VIP cleavage by the two enzymes. Inhibitors of mast cell tryptase and chymase preincubated with the supernatant or with the purified proteases prevented reversal of VIP-induced relaxation. Mast cell proteases did not reverse the tracheal relaxation caused by the nonpeptide adrenergic agonist isoproterenol. These findings show that mast cell proteases tryptase and chymase counteract the smooth muscle relaxant effects of VIP in ferret trachea and suggest a potential role for the mast cell proteases in the modulation of nonadrenergic neural control of airway tone by VIP.
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PMID:Mast cell tryptase and chymase reverse airway smooth muscle relaxation induced by vasoactive intestinal peptide in the ferret. 249 55

Exposure of rat serosal mast cells (RSMC) to chymase, an endogenous secretory granule serine protease, at 37 degrees results in exocytosis, as determined by beta-hexosaminidase release. As the number of RSMC is increased with a set amount of chymase, the net percentage beta-hexosaminidase release decreases linearly, implying a finite set of cellular interactions per chymase unit. Pretreatment of RSMC with trypsin at 37 degrees renders them refractory to subsequent exocytosis mediated by chymase in a dose- and time-dependent fashion, with complete refractiveness occurring by 15 min at 37 degrees with 2.5 micrograms trypsin/ml. Anti-IgE-mediated coupled activation-secretion of RSMC is not affected by the same trypsin pretreatment. When RSMC are pretreated with trypsin (2.5 micrograms/ml) for 0-120 min at 1 degree a progressive loss of sensitivity to activation by chymase at 37 degrees occurs. RSMC susceptibility to chymase-mediated degranulation after trypsin pretreatment can be partially regenerated by culturing the RSMC for about 24 hr in medium at 37 degrees. These findings suggest that a trypsin-sensitive constituent, possibly a receptor or substrate, is necessary for the functional interaction of chymase with RSMC. When added with diisopropyl fluorophosphate (DFP), chymase does not induce RSMC degranulation at 37 degrees. However, if the DFP is removed before addition of chymase at 37 degrees or is added after the chymase-priming event occurs at 1 degree, subsequent degranulation at 37 degrees is not inhibited. Thus, the induction and not the secretion phase is DFP-inhibitable in chymase-induced activation-secretion. In addition, the priming but not the exocytosis phase of chymase-initiated RSMC activation-secretion, which is not dependent on temperature and calcium ion concentration, involves a cellular trypsin-sensitive protein.
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PMID:Modulation of chymase-mediated rat serosal mast cell degranulation by trypsin or diisopropyl fluorophosphate. 252 9

Nucleated cells of human umbilical cord blood were cocultured with mouse skin-derived 3T3 fibroblasts. After 7-8 weeks in culture, when the number of the other hematopoietic cells declined, metachromatic granule-containing mononuclear cells appeared in the cultures, and the number of the cells increased up to 12 weeks. After 11-14 weeks in culture, the metachromatic mononuclear cells comprised a substantial portion of the cultured cells. These cells contained 1.8-2 micrograms of histamine per 10(6) cells and bore receptors for IgE. All of the cells contained tryptase in their granules. Electron microscopic analysis showed that these cells were mature human mast cells, clearly different from the basophilic granulocytes or eosinophils that arise in a variety of circumstances in cord blood cell cultures. Most of the cultured mast cells expressed some granules with regular crystalline arrays and contained both tryptase and chymase, and thus resembled human skin mast cells.
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PMID:Development of human mast cells in vitro. 253 57

Mast cells at immature stages of development were identified in human tissues by electron microscopic techniques. General morphologic criteria of immaturity (such as a high apparent nuclear:cytoplasmic ratio and small cell size), the presence of few granules (those present being smaller than those in mature mast cells) and a lack of features of mast cell activation were used together to determine the level of maturity. Mast cells were identified as being of the T or TC type by immunogold staining with polyclonal rabbit IgG anti-chymase and murine monoclonal anti-tryptase primary antibodies and the appropriate gold-labeled secondary antibodies. Only those cells with tryptase-positive granules were recognized as mast cells. Immature T mast cell granules contained the same characteristic discrete scrolls found in their mature counterparts and all stained positive for tryptase. The presence of trace amounts of chymase in a minority of these granules, as in mature T mast cells, could not be ruled out. The majority of granules in immature TC mast cells had one or more amorphous electron-dense cores rather than the grating and lattice substructures characteristic of granules in mature TC mast cells. Secretory granules in immature TC mast cells stained positively for tryptase and chymase. Occasional immature TC mast cells contained a complete granule or a portion of a granule with the substructure characteristic of mature TC mast cells, favoring the concept that these TC mast cell forms are developmentally related. Essentially all mast cells in foreskin of newborns appeared immature, whereas 10, 5, 10, and 15% of the mast cells in adult lung, foreskin, bowel mucosa and bowel submucosa, respectively, appeared immature. The distribution of T and TC types of immature mast cells seemed to parallel that of the mature mast cell types. These compositional and ultrastructural differences between immature T and TC types of mast cells suggest that from the time granule formation begins, and possibly before this time, each type of human mast cell follows a distinct developmental pathway.
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PMID:Ultrastructural analysis of maturing human T and TC mast cells in situ. 264 87

We tested four synthetic substances for their histochemical value to demonstrate the catalytic activities of chymase or tryptase in mast cells in sections of human gut. Both Suc-Ala-Ala-Phe-4 methoxy-2-naphthylamide (MNA) and N-acetyl-L-methionine-alpha-naphthyl ester (alpha-N-O-Met) reacted with chymase but not tryptase in mast cells. Conversely, D-Val-Leu-Arg-MNA and Z-Ala-Ala-Lys-MNA were hydrolyzed by mast cell tryptase but not chymase. These results were confirmed by use of two inhibitors of chymotrypsin-like activity, chymostatin and Z-Gly-Leu-Phe-chloromethyl ketone (CK) and two inhibitors of trypsin-like activity, Tos-Lys-CK and D-Val-Leu-Arg-CK. Excellent staining reactions were obtained on cryostat sections of unfixed or aldehyde-fixed tissues and on paraffin sections of Carnoy-fixed tissues. For chymase, however, Suc-Ala-Ala-Phe-MNA is preferred on cryostat sections because it is more specific. On paraffin sections alpha-N-O-Met is preferred because other cells are not then stained. For tryptase, Z-Ala-Ala-Lys-MNA was more selective and more specific and is the preferred general purpose substrate on cryostat sections of aldehyde-fixed tissues and for paraffin sections. D-Val-Leu-Arg-MNA is the preferred substrate for cryostat sections of unfixed tissue. Only a limited number of mast cells showed a reaction for chymase, and these occurred mainly in the submucosa. All mast cells, however, gave a reaction for tryptase, and we recommend the use of either substrate for this enzyme for routine detection of mast cells in human tissues. Double staining for the two main mast cell proteases is most conveniently undertaken on paraffin sections of Carnoy-fixed tissues using MNA substrates for tryptase and alpha-N-O-Met for chymase.
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PMID:Enzyme histochemical discrimination between tryptase and chymase in mast cells of human gut. 264 38

The mast cell proteases tryptase and chymase have long been known to constitute one-fifth of the total protein in mast cells. However, their biological functions have not been easy to study because of the difficulty in obtaining sufficient amounts of the enzymes to study their biological functions. Recently, we have been fortunate to have available a permanent line of dog mastocytoma cells to purify both enzymes to homogeneity, and we have used the purified enzymes in two ways. First, in a series of biological studies, we have discovered unique and potent actions of the enzymes that may provide important insights into the pathogenesis of diseases such as asthma and cystic fibrosis. Important biological activities are also likely to exist in other tissues. Because of their structures, mast cell proteases are likely to act in proximity to their sites of release. Thus, the presence and amounts of tryptase and chymase in specific loci may play important roles in tissue responses. In diseases such as asthma and cystic fibrosis, there is evidence that the expression of these mast cell enzymes changes, and these changes have important pathogenetic implications. Second, we have begun to perform structural studies of the enzymes. The recent cloning of tryptase by our group should assist in the better understanding of its functions. Crystallography of the pure proteins should provide further insights and could be the basis of rational development of potent and selective drugs that will inhibit their actions.
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PMID:Roles of mast cell proteases in airways. 266 41


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