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:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The relationship of enzyme structure to substrate specificity for the matrix metalloproteinases interstitial collagenase and stromelysin-2 has been investigated by analysis of the cleavage specificity of recombinant human collagenase-stromelysin-2 hybrid proteins and C terminally truncated collagenase and stromelysin-2. Two series of chimeric proteins were devised by progressive substitution of exon-encoded domains. The recombinant proteins were expressed in COS-7 cells as protein A-fusion proteins and purified on an IgG affinity matrix. Treatment with 4-amino-phenylmercuric acetate released active metalloproteinase of the sizes predicted for the chimeric proteins. Active forms of both the chimeric protein series and the short form enzymes expressed both casein- and gelatin-degrading activities. Like stromelysin, the catalytic activity of stromelysin-2 was contained in the N-terminal domain (encoded by exons 1-5) and was apparently independent of the C-terminal domain (encoded by exons 6-10). Only full-length collagenase displayed a triple helicase (collagenolytic) activity; no combination of N- or C-terminal collagenase domains fused with stromelysin-2 domains had such activity. This suggests that the triple helicase activity is a composite of elements derived from both halves of the collagenase molecule. C terminally truncated collagenase (exons 1-5) and a hybrid of collagenase exons 1-5 and stromelysin-2 exons 6-10 cleaved denatured type I collagen (gelatin) to generate diagnostic peptides in gelatin fingerprint assays. When exon 5 (the exon encoding the zinc-binding domain) was derived from stromelysin-2, the enzyme specificity in the fingerprint assay changed to that of native stromelysin-2. In contrast, when exon 5 was derived from collagenase, the specificity reflected that of the parent enzyme. Our data also suggest that mismatching of exons 2 and 5 destabilizes the enzyme, presumably by altering the geometry of the propeptide-zinc-binding site interaction. We conclude that the loss of triple helicase collagenolytic activity is not accompanied by a shift to the broad specificity characteristic of stromelysin. Rather, the zinc-binding domain confers a distinct cleavage specificity on each metalloproteinase.
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PMID:Role of zinc-binding- and hemopexin domain-encoded sequences in the substrate specificity of collagenase and stromelysin-2 as revealed by chimeric proteins. 846 59

We found that short-term culture medium and homogenate of casein-induced rat peritoneal polymorphonuclear leukocytes (PMN) markedly induced collagenase and prostaglandin E2 (PGE2) production by normal rat synovial cells and these effects were abrogated by anti-(rat interleukin-1 alpha) (IL-1 alpha) polyclonal antibodies. However, collagenase activity and PGE2 induced by recombinant rat IL-1 alpha were less than those induced by rat PMN culture medium. It was also proved by radioimmunoassay that rat PMN culture medium contains a relatively small amount of IL-1 alpha. The introduction of IL-1 alpha-deleted PMN culture medium and recombinant rat IL-1 alpha together into the synovial cell culture system revealed that IL-1 alpha deleted PMN culture medium has a significant enhancing activity on IL-1 alpha-induced synovial cell collagenase and PGE2 production. This new factor, which was shown to be a negatively charged protein of about 80 kDa, may have important roles in connective tissue destruction and chronic inflammation in diseases such as rheumatoid arthritis.
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PMID:A factor derived from polymorphonuclear leukocytes enhances interleukin-1-induced synovial cell collagenase and prostaglandin E2 production in rats. 861 24

The tripeptide compounds, Glu-Arg-Pro-amide (ERPm), D-Pro-Thr-Trp-amide (dPTWm) and thioproline-Thr-Trp (tPTW), were obtained by screening of synthetic peptides for growth-inhibitory activity toward cultured transformed cells. The effects of these peptide compounds on proteases were investigated and the results showed that these compounds enhanced the amidolytic activity of serine proteases despite the fact that each reaction was carried out under optimal conditions. ERPm stimulated the activities of trypsin, chymotrypsin, thrombin, plasmin urokinase and elastase. dPTWm also showed similar effects except that toward chymotrypsin. tPTW elevated the activity only of trypsin, chymotrypsin and thrombin. Stimulation of trypsin activity by these compounds was also confirmed by using casein as a substrate. None of these compounds affected the amidolytic activities of metalloproteinases (MMP-1 and MMP-9), cysteine proteinases (m- and mu-calpains, cathepsin B and papain) or an exopeptidase (leucine aminopeptidase). The activation was at least partly due to the stabilization of the catalytic activity of proteases as well as prevention of autolysis.
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PMID:Enhancement of catalytic activities of serine proteases by tripeptides compounds. 863 1

Following renal ablation, there is marked compensatory renal growth, which is associated with alterations in the activities of renal proteinases. In the present study, rats underwent 5/6 nephrectomy (5/6-NX). Sixteen weeks after surgery, glomeruli and tubules were isolated and proteinase activities were determined using fluorogenic peptidyl substrates. Following 5/6-NX, there was considerable compensatory renal growth resulting in a final weight of 1,923 +/- 46 mg for the remnant kidney as compared to 1,402 +/- 63 mg for the left kidney of SHAM animals. This hypertrophic response was associated with lower activities of tubular cysteine proteinases (cathepsin L & B: -43%; cathepsin B: -61%; cathepsin H: -53%). Significantly reduced activities were also observed for glomerular collagenase (20.2 +/- 6.2 vs. 53.4 +/- 5.7 mU/micrograms DNA) and gelatinase (24.1 +/- 5.0 vs. 130.8 +/- 8.4 mU/micrograms DNA) activities. Protein restriction (5 vs. 20% casein) considerably attenuated compensatory renal growth after surgical ablation (790 +/- 45 vs. 1,923 +/- 46 mg) and partially prevented the fall in tubular cathepsin activities. In terms of glomerular enzymes, protein restriction caused a significant increase in the activity of gelatinase from 24.1 +/- 5.0 to 66.7 +/- 9.2 mU/micrograms DNA, while collagenase remained unchanged. From these data, we conclude that compensatory renal growth is strongly influenced by the amount of protein ingested. It appears that this effect is mediated by modulation of renal proteinase activities.
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PMID:Protein restriction influences glomerular matrix turnover and tubular hypertrophy by modulation of renal proteinase activities. 867 12

Porphyria cutanea tarda is characterized by severe connective tissue damage in sun-exposed skin. The regulated synthesis and degradation of the extracellular matrix by various matrix metalloproteinases (MMPs) determine its amount and composition within the skin. In this study, we therefore asked whether long-wave ultraviolet irradiation (340-450 nm) in conjunction with uroporphyrin I could modulate the synthesis of MMPs with substrate specificities for dermal (collagens I, III, V; proteoglycans) and basement membrane components (collagens IV, VII; fibronectin; laminin) and whether synthesis of the counteracting tissue inhibitor of metalloproteinases is also affected. After irradiation of uroporphyrin-pretreated fibroblasts, specific mRNAs of MMP-1 and MMP-3 increased concomitantly up to 2.7-fold compared with ultraviolet-irradiated cells and up to 10-fold compared with mock-irradiated or uroporphyrin I-treated controls. In contrast, mRNA levels of tissue inhibitor of metalloproteinases remained unaltered. Similar results were obtained by immunoprecipitation. Gelatin and casein zymography revealed increased proteolytic activity of MMP-2 and MMP-3 in blister fluids of patients with porphyria cutanea tarda, indicating that similar events may occur in vivo. Using deuterium oxide as enhancer and sodium azide as quencher of singlet oxygen, we could increase or reduce MMP synthesis, suggesting that singlet oxygen is the major intermediate in the upregulation of MMPs after irradiation of uroporphyrin-pretreated fibroblasts. Taken together, our results show that ultraviolet irradiation alone, and to a greater extent in conjunction with uroporphyrin I, results in an unbalanced synthesis of MMPs that may contribute to the destruction of the dermis and basement membrane, leading to blistering and accelerated photoaging in porphyria cutanea tarda patients.
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PMID:Photosensitization of uroporphyrin augments the ultraviolet A-induced synthesis of matrix metalloproteinases in human dermal fibroblasts. 875 77

Although heart attack is caused by occlusion of a major coronary artery, some patients have occlusion without heart attack because these patients have sufficient collateral circulation to provide an alternate pathway for blood supply to the myocardium at ischemic risk. The growth of new capillary vessels (angiogenesis) and enlargement of preexisting vessels play an important role in the collateral development. We evaluated the hypothesis that extracellular matrix metalloproteinase (MMP) expression is altered in coronary collateral arteries (0.5-1 mm o.d.) isolated from canine hearts 2-4 months after surgical placement of an ameroid occluder around the proximal left circumflex artery (n = 4), during the development of collateral vessels and restructuring new vessels. Histologic studies (hematoxylin and eosin, trichrome, and van Gieson stains) indicated cellular proliferation and increased collagen and elastin content in collateral vessels compared with comparable-sized unoccluded arterial segments of the left anterior descending (LAD) artery. In situ MMP activity of collateral vessels, measured using denatured collagen in the gel matrix, indicated an increase in total MMP activity in the intima of collateral vessels compared with normal LAD vessels. To further identify the type of MMP, tissue homogenates were prepared from collateral and LAD vessels and analyzed by SDS-PAGE zymography. The results suggest induction of gelatinase A and gelatinase B expression in collateral vessels compared with normal LAD tissue, when identical amounts of total protein were loaded onto each lane in the gel. Based on plasminogen-casein zymography, we observed the tissue plasminogen activator level to be increased in collateral vessels. On the basis of immunoblot and mRNA (Northern blot) analyses, we determined that the MMP-1 level was induced in collateral vessels 2 and 4 months after ameroid occlusion. In contrast with MMP-1, the level of TIMP-1 (tissue inhibitor of metelloproteinases) was decreased significantly (p < 0.001) in collateral compared with LAD vessels, suggesting a role for arterial TIMP in anti-angiogenic activity. Collectively, these results suggest that chronic occlusion of a major coronary artery induces upregulation of vascular remodeling mechanisms subserving collateral development. Increased MMP-2 activity in collaterals may be associated with decreased levels of tissue inhibitor of metalloproteinases and fibrous tissue remodeling following angiogenic and (or) adaptive responses of the myocardium to chronic ischemia.
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PMID:Temporal expression of extracellular matrix metalloproteinases and tissue plasminogen activator in the development of collateral vessels in the canine model of coronary occlusion. 896 Mar 89

Activated lamina propria T cells responding to luminal Ags are thought to be important in celiac disease and Crohn's disease, and T cells responding to foreign MHC products are also important in intestinal graft-vs-host disease and intestinal transplant rejection. However, the mechanism(s) by which T cells mediate damage in the gut is not known. We have previously shown that activation of lamina propria T cells by PWM in explant cultures of second trimester human small intestine produces severe tissue injury, with epithelial cell shedding and loss of villi. In this study, we have investigated the role of matrix metalloproteinases in this system. Organ culture supernatants of explants stimulated with PWM showed a 3-fold increase in the concentration of interstitial collagenase and a 10-fold increase in stromelysin-1 compared with control explant culture supernatants. Tissue inhibitors of metalloproteinase-1 and -2 concentrations were unchanged. Increased metalloproteinase enzymatic activity was detected by gelatin and casein zymography. Western blotting revealed the active forms of interstitial collagenase and stromelysin-1 in PWM-stimulated culture supernatants. Up-regulation of mRNA for interstitial collagenase, stromelysin-1, and gelatinase-B was also seen. Nanomolar amounts of recombinant stromelysin-1 added directly to explants produced rapid severe tissue injury. PWM-induced mucosal injury was inhibited by a synthetic peptidomimetic inhibitor of matrix metalloproteinases. Mesenchymal cells isolated from the mucosa of human fetal small intestine produced increased amounts of interstitial collagenase, gelatinase A, and stromelysin-1 when stimulated with IL-1beta or TNF-alpha. These results suggest that T cell activation in the lamina propria results in increased production of matrix metalloproteinases, which by degrading the lamina propria matrix represent a major pathway by which T cells cause injury in the gut.
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PMID:A major role for matrix metalloproteinases in T cell injury in the gut. 902 93

The South American opossum Didelphis marsupialis is known to be highly resistant to snake envenomation. In this paper it is shown that the opossum serum inhibits haemorrhage induced by both Crotalinae and Viperinae venoms. Tested against Bothrops jararaca (jararaca) venom, the antibothropic complex (ABC) isolated from the opossum serum was at least six times more antihaemorrhagic than the commercial antivenom. ABC showed no proteolytic activity by itself and was not hydrolysed by the venom. It inhibited the hydrolysis of casein by B. jararaca venom, but did not inhibit its hydrolytic activities upon N alpha-benzoyl-L-arginine ethyl ester (BAEE) and N alpha-benzoyl-DL-arginine p-nitroanilide (BAPNA). The inhibitor did not interfere with trypsin and bacterial collagenase activities on BAPNA and N-(3-[2-furyl]acryloyl)-Leu-Gly-Pro-Ala (FALGPA), respectively. It reduced chymotrypsin hydrolysis of N-acetyl-L-tyrosine ethyl ester (ATEE) because ABC is also a substrate for this enzyme. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, B. jararaca venom preferentially degraded fibrinogen A alpha-chain and fibrin alpha-chain. Tested on extracellular matrix proteins, the venom hydrolysed collagen IV, gelatins I and V, laminin and fibronectin, besides depolimerizing collagen I alpha-chain dimers. Fibrillar collagen V was not digested. These hydrolyses were inhibited by ABC and by EDTA. Our results show that the antibothropic complex is a venom metalloproteinase inhibitor, which could, at least partially, account for its antihaemorrhagic activity. Electrophoretic evidence indicated non-covalent complex formation between the antihaemorrhagic factor and component(s) of B. jararaca venom.
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PMID:Inhibitory properties of the antibothropic complex from the South American opossum (Didelphis marsupialis) serum. 924 80

The production of proteolytic enzymes by 10 Clostridium difficile isolates of varying toxigenicity and clinical origin was studied to determine if all isolates secreted proteases. Different protease substrates were studied: gelatin, collagen, phenylazobenzyloxycarbonyl-leucyl-glycyl-L-prolyl-D-arginine (Pz-peptide), casein, azocasein, and azocoll. All isolates degraded gelatin, collagen, and azocoll. The supernatants of all isolates contained an enzyme capable of attacking gelatin incorporated in a polyacrylamide gel (zymograms) and forming two closely spaced lytic bands with an estimated molecular mass of 35-40 kDa. Polyclonal antibodies, produced against the C. difficile gelatinase, revealed in Western blots a 35-kDa protein in the culture supernatants of all C. difficile isolates. In the same manner, Clostridium perfringens collagenase polyclonal antibodies detected a 120-kDa protein in the culture supernatants of all isolates; this suggests that at least two proteases may exist in C. difficile. The protease activities of the 10 strains examined did not seem strikingly different quantitatively but were in general weak and their role in pathogenicity is suspect.
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PMID:Protease activity of Clostridium difficile strains. 954 17

We cloned a novel matrix metalloproteinase (MMP) called CMMP from cultured primary chicken embryo fibroblasts. The cDNA-derived CMMP sequence contains 472 amino acids including a putative 19-residue signal peptide and a unique cysteine in the catalytic domain, an insertion in a sequence motif that binds the structural (noncatalytic) zinc of MMPs. Strikingly, a homologously inserted cysteine is also found in Xenopus XMMP and human MMP19, two recently cloned novel members of the MMP family. Phylogenetic analysis suggest that XMMP and MMP19 represent founding members of the MMP family, whereas CMMP is related to collagenase MMPs. Bacterially produced recombinant CMMP (without the amino-terminal inhibition domain), which was autoproteolyzed at the carboxyl-terminal domain, digested casein and gelatin. As shown by Northern blotting, CMMP mRNA of 1.8 kilobase pairs was constitutively expressed in cultured primary chicken embryo fibroblasts and up-regulated by tumor necrosis factor-alpha and the phorbol ester 12-O-tetradecanoylphorbol-13-acetate, but it was not regulated by interleukin-1, basic fibroblast growth factor, or retinoic acid. CMMP mRNA of 1.8 kb was also detected in the head and body of 8-day-old chicken embryos and dramatically up-regulated in 9-day-old embryos.
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PMID:Cloning and characterization of a novel matrix metalloproteinase (MMP), CMMP, from chicken embryo fibroblasts. CMMP, Xenopus XMMP, and human MMP19 have a conserved unique cysteine in the catalytic domain. 965 95


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