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 peptide derivative N alpha-(2,4-dinitrophenyl)-L-prolyl-L-leucyl-glycyl-L-prolyl-L-tryptophanyl-D- lysine (Dnp-Pro-Leu-Gly-Pro-Trp-D-Lys) has been found to be a convenient substrate for the assay of clostridial collagenase and Pz-peptidase. The substrate shows a 25-fold enhancement of fluorescence (gamma ex. 283 nm, lambda em. 350 nm) following hydrolysis of the Leu2-Gly3 peptide bond. The value of Km for clostridial collagenase was 17 microM. The substrate for the first time makes possible continuous fluorimetric assays for Pz-peptidase and clostridial collagenase.
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PMID:A continuous fluorimetric assay for clostridial collagenase and Pz-peptidase activity. 254 53

Pz-peptidase was purified from rabbit muscle by acid precipitation of tissue homogenate followed by cation- and anion-exchange chromatography, gel chromatography, and immunoadsorption. In analytical gel chromatography, one single peak of protein with corresponding Pz-peptidase activity was obtained. The enzyme had an apparent Mr of 74,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and was eluted at pH 4.8 in chromatofocusing. No metals were detectable in the protein by neutron activation analysis. Purified Pz-peptidase hydrolyzed Dnp-Pro-Leu-Gly-Pro-Trp-D-Lys (Km 7.2 microM) most effectively in the presence of 5 mM 2-mercaptoethanol and 10 mM CaCl2. No inhibition was observed with inhibitors of serine proteinases, aspartic proteinases, or metalloproteinases, apart from some nonspecific reversible inhibition by 1,10-phenanthroline. The activation by Ca2+ was reversed by EDTA. The enzyme was not inhibited by E-64, cystatin, or leupeptin, but was irreversibly inactivated by iodoacetate, iodoacetamide, and N-ethylmaleimide. It was therefore concluded that rabbit muscle Pz-peptidase is not a typical member of any of the four recognized catalytic classes of proteinases, but may be an atypical cysteine endopeptidase. The peptidase was not bound by alpha 2-macroglobulin. No hydrolysis of gelatin or fibronectin by the enzyme was detected, nor was there any activation of latent collagenase.
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PMID:Purification and characterization of Pz-peptidase from rabbit muscle. 267 41

Human skin, maintained in serum-free organ culture, secretes a neutral metalloendopeptidase which is remarkably specific for gelatin. Because the product peptides from the action of collagenase on collagen become denatured into random coil polypeptides of 25000 and 75000 daltons at physiological temperature, it is thought that this "gelatinase" is the second, and possibly the only other enzyme in the pathway of extracellular collagen degradation. New types of high-performance liquid chromatography (HPLC) columns have enabled us to improve the yields of active gelatinase from skin culture medium. Raw medium, which has been dialyzed and lyophilized, is fractionated with ammonium sulfate, and applied to Pharmacia Blue Sepharose in a batch step. The 0.4 M sodium chloride eluate is then subjected to gel filtration on Sephacryl S-200, followed by gradient elution from Amicon Green Sepharose. The fractions with gelatinolytic activity are applied to a Bio-Rad TSK-Phenyl-5PW HPLC column for mild hydrophobic chromatography with a gradient of decreasing ammonium sulfate concentration. In the final step, the enzyme is applied to a Pharmacia Mono-Q FPLC column and eluted with a gradient of sodium chloride. At this point, the enzyme appears as two bands, corresponding to enzymatic activity zymograms on sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
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PMID:Purification of gelatin-specific neutral protease from human skin by conventional and high-performance liquid chromatography. 299 26

A third metalloendopeptidase activity, gelatinase, has been completely separated from the collagenase and proteoglycanase activities of rabbit bone culture medium. Although the proteinase could not be purified to homogeneity in large amounts, it was possible to obtain accurate molecular weight values and activity after electrophoresis on non-reduced SDS/polyacrylamide gels. The latent form had an Mr of 65 000 which could be activated with 4-aminophenylmercuric acetate, APMA, to a form of Mr 61 000; under reducing conditions the latent and active forms had Mr of 72 000 and 65 000, respectively. Trypsin was a very poor activator of the latent enzyme. Gelatinase degraded gelatins derived from the interstitial collagens and it also had low activity on native types IV and V collagen and on insoluble elastin. Gelatinase acted synergistically with collagenase in degrading insoluble interstitial collagen. The specific mammalian tissue inhibitor of metalloproteinases inhibited gelatinase by forming a stable inactive complex. Comparison of the properties of gelatinase with those of collagenase and proteoglycanase suggest that the three proteinases form a family which together are capable of degrading all the major macromolecules of connective tissue matrices.
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PMID:Purification and characterization of a bone metalloproteinase that degrades gelatin and types IV and V collagen. 299 41

Changes in cell shape are postulated to modulate gene expression during differentiation of a number of cell types, including rabbit synovial fibroblasts, which are inducible for expression of the zymogen form of the metalloendopeptidase, collagenase. In the work presented here, fibroblasts cultured on and within hydrated collagen gels were allowed to contract by release of the gels from the sides of the culture dish. Within 24 h of cell release, synthesis and secretion of procollagenase was initiated in the absence of any chemical manipulation. Fibroblasts grown in and on collagen also responded to 12-O-tetradecanoylphorbol-13-acetate and cytochalasin B with morphologic change and induced procollagenase. However, colchicine, which altered morphology to varying degrees in cells on plastic, on collagen, and within collagen gels, did not induce procollagenase expression. In all cases, the enzyme was induced only after reorganization of polymerized actin, rather than after a change in cellular morphology per se. As a first approach to identifying other aspects of the stimulated phenotype that could affect collagen turnover, the expression of collagen and endogenous metalloproteinase inhibitors in relation to procollagenase secretion was investigated. Collagen secretion by fibroblasts decreased when procollagenase secretion was induced by the pharmacologic agents, but not when cells were stimulated by contraction on or within collagen gels. The expression of two endogenous inhibitors was not coordinately regulated with induction of procollagenase. Therefore, the extracellular matrix and the cellular actin cytoskeleton may transduce signals that modulate the tissue remodeling phenotype of fibroblasts.
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PMID:Reorganization of polymerized actin: a possible trigger for induction of procollagenase in fibroblasts cultured in and on collagen gels. 301 94

A neutral protease with a marked specificity for gelatin as a protein substrate has been purified to homogeneity from medium of human skin in serum-free explant culture. The pH optimum of this gelatinase is between 7.0 and 7.5 with little or no activity displayed below pH 5. Inhibition by EDTA, ethylene glycol bis(beta-amino-ethyl ether)N,N,N',N'-tetraacetic acid (EGTA), and 1,10-phenanthroline suggest that the enzyme is a metalloendopeptidase. Calcium concentration-dependent inhibition of the enzyme by EGTA and EDTA suggest further that there is a requirement for extrinsic calcium. Indeed, removal of calcium and reduces enzyme activity, and subsequent addition of calcium restores full activity. The gelatinase is not inhibited by serine protease inhibitors but is inhibited by cysteine, dithiothreitol, and beta-mercaptoethanol. It is also inhibited by a macromolecular inhibitor of collagenase which has been purified from human skin fibroblasts. The apparent molecular weight of this enzyme, as determined by gel filtration is 120,000-150,000. The enzyme is a glycoprotein, as indicated by staining with periodic acid-Schiff reagent and by its affinity for lectins. Human skin gelatinase shows little or no reactivity toward common protein substrates, such as hemoglobin or casein, and does not cleave helical collagen. Two sites of cleavage in the sequence of gelatin, Gly-Ile and Gly-Leu, have been positively identified using synthetic substrates and tryptic peptides of collagen.
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PMID:Purification and properties of a gelatin-specific neutral protease from human skin. 626 Aug 9

Cells derived from isolated glomerular tufts of rats were studied in primary tissue culture after the removal of epithelial cells by collagenase treatment. The cultured cells, fusiform or stellate in shape, grew readily over a 12-day period. Immunofluorescence staining was positive for myosin and fibronectin, while negative for Factor VIII, suggesting that the outgrowing cells were derived from the glomerular mesangium. In serum-free culture, these cells produced neutral proteinase activity that occurred as a latent trypsin-activable form (apparent molecular weight range, 78,000 to 100,000 daltons) and in an active form (44,000 to 58,000 daltons). Neutral proteinase activity was inhibited by EDTA and by cysteine, and exhibited a pH optimum of 7.2 to 7.8, characteristic of an extracellularly active metalloendopeptidase. The culture supernate which contained the neutral proteinase activity was capable of degrading purified rat glomerular basement membrane. The release of hydroxyproline-containing fragments from the basement membrane indicated that degradation of the type IV collagen component of the basement membrane was occurring. These findings suggest that the neutral proteinase activity generated by mesangium-derived cells may play a role in the physiologic turnover of glomerular structural proteins in vivo.
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PMID:Neutral proteinase activity produced in vitro by cells of the glomerular mesangium. 640 73

Thimet oligopeptidase (EC 3.4.24.15) is a thiol-dependent metallo-endopeptidase also known as Pz-peptidase, collagenase-like peptidase, endooligopeptidase A, soluble metallo-endopeptidase and endopeptidase 24.15. The enzyme is closely related to the yeast proteinase yscD. Thimet oligopeptidase (M(r) 74000) is widely distributed in animals and plants. In rat liver it exists in a cytoplasmic and mitochondrial form; a membrane-bound form of the enzyme was discovered in rat brain. Thimet oligopeptidase hydrolyses small peptides but does not act on proteins. In rat brain thimet oligopeptidase is involved in the generation of enkephalins and inactivation of bioactive peptides and experiments with yeast provided good evidence that the enzyme is involved in the late stages of cytoplasmatic protein degradation.
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PMID:Thimet oligopeptidase--a review of a thiol dependent metallo-endopeptidase also known as Pz-peptidase endopeptidase 24.15 and endo-oligopeptidase. 847 Nov 82

The Clostridium histolyticum 116-kDa collagenase consists of four segments, S1, S2a, S2b, and S3. A 98-kDa gelatinase, which can degrade denatured but not native collagen, lacks the C-terminal fragment containing a part of S2b and S3. In this paper we have investigated the function of the C-terminal segments using recombinant proteins. Full-length collagenase degraded both native type I collagen and a synthetic substrate, Pz-peptide, while an 88-kDa protein containing only S1 and S2a (S1S2a) degraded only Pz-peptide. Unlike the full-length enzyme, S1S2a did not bind to insoluble type I collagen. To determine the molecular determinant of collagen binding activity, various C-terminal regions were fused to the C terminus of glutathione S-transferase. S3 as well as S2bS3 conferred collagen binding. However, a glutathione S-transferase fusion protein with a region shorter than S3 exhibited reduced collagen binding activity. S3 liberated from the fusion protein also showed collagen binding activity, but not S2aS2b or S2b. S1 had 100% of the Pz-peptidase activity but only 5% of the collagenolytic activity of the full-length collagenase. These results indicate that S1 and S3 are the catalytic and binding domains, respectively, and that S2a and S2b form an interdomain structure.
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PMID:A study of the collagen-binding domain of a 116-kDa Clostridium histolyticum collagenase. 945 93

Deterioration of the aortic wall resulting in formation of aneurysm may be evoked by increased activity of elastases, collagenases and lysosomal proteases. These enzymes come from macrophages and neutrophil granulocytes which are elements of the inflammatory reaction accompanying aneurysm. These cells may also come from parietal thrombus in the aneurysm lumen. The aim of this work was to determine activity of elastase, cathepsin G, collagenase-like Pz-peptidase and cathepsins A, B, C, D and E in the parietal thrombus of aortic aneurysm. The thrombus was obtained from the lumen of the aortic aneurysm of six patients during operation. Protease activities were determined using specific substrates at optimum pH. Retracted blood clot was a comparative material. The thrombus of aortic aneurysm showed two-five fold higher activity of elastases, collagenase-like Pz-peptidase and cathepsins A, D and G in comparison to the blood clot (P < 0.001). However, activity of cathepsins B, C and E in the thrombus was only slightly higher (P < 0.05). Prolonged effect of proteases coming from parietal thrombus on the aneurysm wall could evoke marked degradation of fibrillar proteins resulting in increase of aneurysm.
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PMID:Activities of proteases in parietal thrombus of aortic aneurysm. 956 32


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