Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.37 (neutrophil elastase)
 4,078 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The precursor of matrix metalloproteinase 9 (proMMP-9), also known as '92 kDa progelatinase/type IV procollagenase', was purified from the conditioned medium of U937 monocytic leukaemia and HT1080 fibrosarcoma cell lines stimulated with phorbol 12-myristate 13-acetate. ProMMP-9 in these culture media is non-covalently complexed with the 29 kDa tissue inhibitor of metalloproteinases (TIMP), but free proMMP-9 was separated from the TIMP-proMMP-9 complex by chromatography on Green A Dyematrex gel. The final product was homogeneous on SDS/PAGE, with a molecular mass of 88 kDa without reduction and 92 kDa with reduction. Treatment of proMMP-9 with 4-aminophenylmercuric acetate converted the 88 kDa precursor into 80 kDa and 68 kDa forms. Gelatin-containing zymographic analysis showed zones of lysis associated with all three species. However, only the 68 kDa species was shown to be catalytically active by its ability to bind to alpha 2-macroglobulin. In the presence of an equimolar amount of TIMP, only the 80 kDa species was generated by treatment with 4-aminophenylmercuric acetate, but no enzyme activity was detected. This indicates that TIMP binds to the 80 kDa intermediate and inhibits the generation of the active 68 kDa species. Eight endopeptidases (trypsin, chymotrypsin, plasmin, plasma kallikrein, thrombin, cathepsin G, neutrophil elastase and thermolysin) were tested for their ability to activate proMMP-9. Of them, trypsin was the most effective activator of proMMP-9. Only partial activation (10-30%) was observed with plasmin, cathepsin G and chymotrypsin. The active forms generated by trypsin were identified as 80 kDa, 74 kDa and 66 kDa by their abilities to bind to alpha 2-macroglobulin. In the presence of an equimolar amount of TIMP, proMMP-9 was also converted into the same molecular-mass species by trypsin, but they were not proteolytically active. This suggests activated MMP-9 is inhibited by TIMP. Activated MMP-9 digested gelatin, type-V collagen, reduced carboxymethylated transferrin and, to a lesser extent, type-IV collagen and laminin A chain. The specific activity against gelatin was estimated to be 15,000 units/mg (1 unit = 1 microgram of gelatin degraded/min at 37 degrees C) by titration with alpha 2-macroglobulin. Comparative studies on digestion of gelatin and collagen types IV and V by MMP-9 and MMP-2 indicated that both enzymes degrade these substrates into similar fragments. However, the susceptibilities of laminin, fibronectin and reduced carboxymethylated transferrin to these two MMPs were sufficiently different to indicate differences in substrate specificities between these two closely related proteinases.
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PMID:Purification and characterization of matrix metalloproteinase 9 from U937 monocytic leukaemia and HT1080 fibrosarcoma cells. 137 48

Matrix metalloproteinase 9 (MMP-9) has been purified as an inactive zymogen of M(r) 92,000 (proMMP-9) from the culture medium of HT 1080 human fibrosarcoma cells. The NH2-terminal sequence of proMMP-9 is Ala-Pro-Arg-Gln-Arg-Gln-Ser-Thr-Leu-Val-Leu-Phe-Pro, which is identical to that of the 92-kDa type IV collagenase/gelatinase. The zymogen can be activated by 4-aminophenylmercuric acetate, yielding an intermediate form of M(r) 83,000 and an active species of M(r) 67,000, the second of which has a new NH2 terminus of Met-Arg-Thr-Pro-Arg-(Cys)-Gly-Val-Pro-Asp-Leu-Gly-Arg-Phe-Gln-Thr- Phe-Glu. Immunoblot analyses demonstrate that this activation process is achieved by sequential processing of both NH2- and COOH-terminal peptides. TIMP-1 complexed with proMMP-9 inhibits the conversion of the intermediate form to the active species of M(r) 67,000. The proenzyme is fully activated by cathepsin G, trypsin, alpha-chymotrypsin, and MMP-3 (stromelysin 1) but not by plasmin, leukocyte elastase, plasma kallikrein, thrombin, or MMP-1 (tissue collagenase). During the activation by MMP-3, proMMP-9 is converted to an active species of M(r) 64,000 that lacks both NH2- and COOH-terminal peptides. In addition, HOCl partially activates the zymogen by reacting with an intermediate species of M(r) 83,000. The enzyme degrades type I gelatin rapidly and also cleaves native collagens including alpha 2 chain of type I collagen, collagen types III, IV, and V at undenaturing temperatures. These results indicate that MMP-9 has different activation mechanisms and substrate specificity from those of MMP-2 (72-kDa gelatinase/type IV collagenase).
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PMID:Matrix metalloproteinase 9 (92-kDa gelatinase/type IV collagenase) from HT 1080 human fibrosarcoma cells. Purification and activation of the precursor and enzymic properties. 140 Apr 81

The ability of human neutrophil elastase and cathepsin G to activate matrix metalloproteinase 3 (MMP-3 = stromelysin) and MMP-2 ('gelatinase') purified from human rheumatoid synovial fibroblasts in culture was examined. The zymogen of MMP-3 (proMMP-3) was activated to full activity with elastase and cathepsin G by limited proteolysis of the molecule into two active forms of Mr approximately 45,000 and Mr approximately 25,000. In contrast, proMMP-2 was not activated at all by these neutrophil serine proteinases, although it was degraded into small fragments. These data suggest that neutrophil elastase and cathepsin G may play an important role in the activation of proMMP-3 in vivo in various inflammatory conditions, but proMMP-2 may be activated in different ways.
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PMID:Activation of matrix metalloproteinase 3 (stromelysin) and matrix metalloproteinase 2 ('gelatinase') by human neutrophil elastase and cathepsin G. 254 55

The degradation of tenascin purified from human melanoma cells was examined by treatment with matrix metalloproteinases (MMPs) and serine proteinases. Among eight different types of proteinases examined, MMP-1, -3, and -7, cathepsin G and leukocyte elastase could digest tenascin, but MMP-2, MMP-9 and thrombin did not. This suggests that tenascin may be readily catabolized by extracellular matrix-degrading proteinases found in the pathophysiological conditions.
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PMID:Susceptibility of tenascin to degradation by matrix metalloproteinases and serine proteinases. 752 86

Matrix metalloproteinase 7 (MMP-7) has been purified as an inactive zymogen of M(r) 28,000 (proMMP-7) from the culture medium of CaR-1 human rectal carcinoma cells. The NH2-terminal sequence of proMMP-7 is Lys-Pro-Lys-Pro-Gln-Glu, which is identical to that of matrilysin. The zymogen is activated by 4-aminophenylmercuric acetate (APMA), yielding an intermediate form of M(r) 21,000 and an active species of M(r) 19,000 which shows the new NH2-terminal sequence of Tyr78-Ser-Leu-Phe-Pro-Asn-Ser. Although trypsin fully activates the zymogen, the activation rate by plasmin or leukocyte elastase is confined to approximately 50%. ProMMP-7 can be activated by MMP-3 (stromelysin 1) to its full activity in a single-step mechanism and generates the same NH2 terminus obtained by APMA activation, whereas MMP-1 (tissue collagenase), MMP-2 (gelatinase A), and MMP-9 (gelatinase B) do not have such an effect. On the other hand, proMMP-1 is activated by MMP-7 to an activity similar to that obtained by APMA and the activation by MMP-7 is enhanced up to approximately 6.5 fold in the presence of APMA. This enhanced activity is donated by specific cleavage at the Gln80-Phe81 bond of proMMP-1. MMP-7 can also activate proMMP-9 up to approximately 50% of the full activity with a new NH2 terminus of Leu16-Arg-Thr-(Asn)-Leu. Incubation of proMMP-2 or proMMP-3 with MMP-7 results in no activation of these proMMPs. MMP-7 degrades type IV collagen, laminin-1, fibronectin, proteoglycan, type I gelatin, and insoluble elastin. These results suggest that in vivo MMP-7 may play a role in degradation of extracellular matrix macromolecules in concert with MMP-1, -3, and -9 under pathological conditions.
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PMID:Matrix metalloproteinase 7 (matrilysin) from human rectal carcinoma cells. Activation of the precursor, interaction with other matrix metalloproteinases and enzymic properties. 789 11

We present the first direct biochemical evidence for the turnover of intact type VI collagen microfibrils. Matrix-degrading enzymes of the serine proteinase class, including rat mast cell chymases I and II, human mast cell tryptase, neutrophil elastase, cathepsin G and trypsin, were able to catabolize intact type VI collagen microfibrils isolated from foetal bovine skin and metabolically labelled intact type VI collagen immunoprecipitated from fibroblast culture medium. By contrast, intact type VI collagen was not degraded by the human matrix metalloproteinases, MMP-1, MMP-2, MMP-3 and MMP-9. These data have important implications for the stability of type VI collagen in connective tissues and highlight the potential role of serine proteinases both in normal type VI collagen turnover and in inflammatory conditions characterized by matrix degradation.
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PMID:Catabolism of intact type VI collagen microfibrils: susceptibility to degradation by serine proteinases. 846

The aim of this study was to investigate the extracellular degrading proteolytic cascade proteins referred to as matrix metalloproteinase-1 (MMP-1), MMP-2, MMP-9, membrane-type matrix metalloproteinase-1 (MT1-MMP), tissue inhibitors of matrix metalloproteinase-1 (TIMP-1), TIMP-2, neutrophil elastase, and alpha1-antitrypsin in human pulmonary emphysema. Localization of MMP-1, MMP-2, MMP-8, MMP-9, MT1-MMP, TIMP-1, and TIMP-2 was verified by immunohistochemical analysis. The results of our study indicated that the immunoreactivity of MMP-1, MMP-8, MMP-9, and TIMP-1 was absent, whereas MT1-MMP and MMP-2 were mainly observed in pneumocytes, fibroblasts, and alveolar macrophages. Although MT1-MMP and MMP-2 were observed both in emphysematous and normal lung tissue, these immunoreactivities were intense in the emphysematous samples. The presence of MMP-1, MMP-2, MMP-9, TIMP-1, and TIMP-2 was confirmed at mRNA level by reverse transcription-PCR analysis and enzyme immunoassay (EIA). However, the only statistical difference that was observed was in MMP-2 and MMP-9 (MMP-2: emphysematous samples, 19.1+/-2.1 versus control samples, 5.2+/-0.60 microg/g protein, p < 0.05; MMP-9: emphysematous samples, 18.4+/-5.6 versus control samples, 8.1+/-2.7 microg/g protein, p < 0.05). Results of the neutrophil elastase as analyzed by EIA, and alpha1-antitrypsin levels as detected by laser nephelometric immunoassay, indicated no statistical difference between the emphysematous and control groups. In addition to the presence of mRNA levels, the level of MT1-MMP according to immunoblot analysis increased in the emphysematous samples. Gelatin zymographic analysis confirmed the presence of both pro and active forms of MMP-2, and the increased ratio of the active form of MMP-2 in emphysematous samples (25.9%+/-2.0% versus 11.2%+/-3.3%, p < 0.05), indicated in situ activation of MMP-2 by MT1-MMP. Elastin zymographic analysis showed elastolytic activity by MMP-2 and MMP-9 but not the reported band of macrophage metalloelastase (MMP-12). The data suggest that the MT1-MMP/MMP-2/TIMP-2 system plays a significant role in the MMP-mediated extracellular matrix degradation and tissue remodeling of emphysematous lungs, and thus may contribute to the weakening of lung parenchyma and lead to the formation of emphysema.
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PMID:Matrix metalloproteinase-mediated extracellular matrix protein degradation in human pulmonary emphysema. 975 52

Cervical ripening is analogous to an inflammatory reaction characterized by an influx of inflammatory cells and an increase in inflammatory mediators. The anti-gestogen mifepristone is highly effective in inducing cervical ripening in women throughout gestation. However, its mechanism of action is largely unknown. The aim of the study was to investigate the effect of in-vivo administration of mifepristone on inflammatory cells and mediators in the cervix. Cervical biopsies were taken from women undergoing a first trimester termination of pregnancy at 0, 6, 12, 24 and 36 h (n = 6 per group) after mifepristone administration. Biopsies were fixed for immunohistochemistry and also cultured for subsequent analysis of culture media by radioimmunoassay or enzyme-linked immunosorbent assay. After administration of mifepristone (6-24 h), there was an increase in immunostaining for leukocyte common antigen (CD45), neutrophil elastase, monocytes (CD68), and matrix metalloproteinases (MMP)-1, -8 and -9. Immunostaining for MMP-2 and tissue inhibitor of metalloproteinases (TIMP)-1, -2 and -4 were unaffected by mifepristone treatment. Secretion of monocyte chemotactic protein (MCP-1) was significantly (P < 0.05) increased from biopsies taken 6-24 h after mifepristone administration. Cervical biopsies also released interleukin-8 (IL-8), prostaglandin (PG) E(2), PGF(2alpha) and prostaglandin metabolites (PGEM and PGFM) although their secretion was unaffected by mifepristone treatment. This study suggests that mifepristone may, in part, effect cervical ripening by modulating the influx of inflammatory cells into the cervix, up-regulating MMP expression and inducing chemokine secretion by cervical tissue.
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PMID:The effect of mifepristone administration on leukocyte populations, matrix metalloproteinases and inflammatory mediators in the first trimester cervix. 1082 72

Proteinases are important at several phases of physiological and pathological inflammation, mediating cellular infiltration, cytokine activation, tissue damage, remodeling, and repair. However, little is known of their role in the pathogenesis of inflammatory bowel disease. The aim of this study was to assess the role of tissue proteases in a mouse model of colitis. Proteolytic activity was analyzed, using gel and in situ zymography, in colonic tissues from severe combined immunodeficient mice with colitis induced by transfer of CD4(+) T lymphocytes. Serine proteinase levels increased in colitic tissue, with major species of 23 kd, 30 kd, and 45 kd. Co-migration and inhibition studies indicated that the 23-kd proteinase was pancreatic trypsin and that the 30-kd species was neutrophil elastase. Matrix metalloproteinase (MMP)-9 expression, and MMP-2 and MMP-9 activation, was elevated in colitic tissues. Proteinase levels followed a decreasing concentration gradient from proximal to distal colon. Proteolysis was localized to infiltrating leukocytes in diseased severe combined immunodeficient mice. Transmural inflammation was associated with serine proteinase and MMP activity in overlying epithelium and with marked subepithelial proteolytic activity. The results demonstrate a clear elevation in the levels and activation of proteases in colitis, potentially contributing to disease progression through loss of epithelial barrier function.
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PMID:The role of up-regulated serine proteases and matrix metalloproteinases in the pathogenesis of a murine model of colitis. 1110 65

Ulcerative keratitis is a common and potentially blinding ocular disease of horses, capable of progressing to corneal perforation in as little as 24 h. This rapid stromal degeneration is mediated in part by exogenous and endogenous proteinases. We measured and compared the concentrations of two matrix metalloproteinases (MMP-2 and MMP-9) and a serine proteinase (neutrophil elastase) present in the precorneal tear film of normal horses and horses with rapidly progressing ulcerative keratitis. Precorneal tear film samples were collected from 23 ulcerated and 21 unaffected eyes of 23 horses with unilateral ulcerative keratitis, and from 33 normal eyes of 17 control horses. MMP-2, MMP-9, and neutrophil elastase were identified by casein and gelatin zymography and quantified by computerized image analysis. Median MMP-9 levels were significantly higher in the precorneal tear film of young control horses vs. older control horses (P = 0.005). Median MMP-2, MMP-9, and neutrophil elastase levels were significantly higher in the precorneal tear film of ulcerated eyes when compared to age-matched normal controls (P = 0.004, P = 0.001, and P = 0.012, respectively). Median MMP-2 levels were also significantly higher in the precorneal tear film of contralateral eyes of affected horses when compared to age-matched normal controls (P = 0.004). No significant differences in median proteinase levels were detected between 'sterile' ulcers and those from which bacteria or mixed infections (bacteria and fungi) were isolated. However, median MMP-2 and neutrophil elastase levels were significantly higher in the precorneal tear film of eyes with 'sterile' ulcers when compared with ulcerated eyes from which fungi were isolated (P < 0.05). The results of this study support the use of topical antiproteinase therapy which targets both MMPs and serine proteinases in progressive equine ulcerative keratitis.
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PMID:Evaluation of tear film proteinases in horses with ulcerative keratitis. 1139 92


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