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 N-terminal propeptide of type III procollagen was purified from human ascitic fluid by using (NH4)2SO4 precipitation, DEAE-Sephacel chromatography at pH 8.6, Sephacryl S-300 chromatography and another DEAE-Sephacel chromatography at pH 4.5. The Mr of the human peptide was about 42 000, which corresponds in size to the propeptide released by the specific N-proteinase during the extracellular processing of collagen. Bacterial-collagenase digestion of the human peptide produced three fragments, which could be separated on a Bio-Gel P-10 column. The human propeptide and its collagenase-derived fragments, an N-terminal non-collagenous domain Col 1, a C-terminal non-helical domain Col 2 and a collagenous domain Col 3, resembled those derived from the N-terminal segment of bovine type III procollagen in their amino acid composition. The human peptide was found to contain sulphate, which may explain its extremely low isoelectric point (3.1). Antibodies against the human N-terminal propeptide reacted similarly with both the purified human peptide and a corresponding segment of bovine type III procollagen. The human propeptide could be used in developing radioimmunoassays for monitoring fibrotic processes.
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PMID:Purification and characterization of the N-terminal propeptide of human type III procollagen. 408 23

A unique low molecular weight collagen that was highly resistant to proteolytic degradation was originally isolated from fetal calf ligamentum nuchae fibroblasts and hence termed FCL-1 [Sage, H., Mecham, R., Johnson, C., & Bornstein, P. (1983) J. Cell Biol. 97, 1933-1938]. The differential expression of this protein was studied as a function both of fetal (donor) age and of subcultivation in vitro. Concomitant isolation, subculture, and metabolic radiolabeling experiments performed on cell strains from fetal calf ligament (FCL) and fetal bovine skin (FBS) representing different gestational ages (85-270 days in utero) showed that (a) FCL-1 was synthesized preferentially by fibroblasts from younger animals and (b) expression of FCL-1 diminished as a function of increased passage in culture. Levels of FCL-1, measured as percent of total radiolabeled culture medium protein that precipitated in a concentration range of 20-50% ammonium sulfate, ranged from 22% in FCL 85 cells to 7.7% in FCL 270 (term) cells. FBS fibroblasts at passages 6-10 secreted from 13% to 6% FCL-1, respectively. When cells from an 85-day fetal ligament were allowed to accumulate copious extracellular matrix in vitro, the production of FCL-1 was increased to 32%. FCL-1 was not immunoreactive with polyclonal antibodies directed toward most of the sequences of the interstitial type I and type III procollagens. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the apparent molecular weight of FCL-1 was 13 000 (on the basis of collagen peptide standards) and approximately 30 000 (on the basis of globular protein standards). Incubation with bacterial collagenase produced a stable cleavage product of Mr 8000 (by collagen standards) or 17 000 (by globular standards). In contrast, pepsin removed a small peptide of approximately 1000-2000 in molecular weight from FCL-1, and a gradual but progressive proteolysis of the collagen was observed over a period of 1-6 h. Pulse-chase studies revealed a secretion time of approximately 60 min for FCL-1, without the appearance of any processed, intermediate forms. These studies confirm that FCL-1 represents a novel member of the collagen gene family that manifests differential expression as a function of development.
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PMID:Low molecular weight fibroblast collagen: structure, secretion, and differential expression as a function of fetal and cellular age. 408 89

1. A method is described for extracting separately mitochondrial and extramitochondrial enzymes from fat-cells prepared by collagenase digestion from rat epididymal fat-pads. The following distribution of enzymes has been observed (with the total activities of the enzymes as units/mg of fat-cell DNA at 25 degrees C given in parenthesis). Exclusively mitochondrial enzymes: glutamate dehydrogenase (1.8), NAD-isocitrate dehydrogenase (0.5), citrate synthase (5.2), pyruvate carboxylase (3.0); exclusively extramitochondrial enzymes: glucose 6-phosphate dehydrogenase (5.8), 6-phosphogluconate dehydrogenase (5.2), NADP-malate dehydrogenase (11.0), ATP-citrate lyase (5.1); enzymes present in both mitochondrial and extramitochondrial compartments: NADP-isocitrate dehydrogenase (3.7), NAD-malate dehydrogenase (330), aconitate hydratase (1.1), carnitine acetyltransferase (0.4), acetyl-CoA synthetase (1.0), aspartate aminotransferase (1.7), alanine aminotransferase (6.1). The mean DNA content of eight preparations of fat-cells was 109mug/g dry weight of cells. 2. Mitochondria showing respiratory control ratios of 3-6 with pyruvate, about 3 with succinate and P/O ratios of approaching 3 and 2 respectively have been isolated from fat-cells. From studies of rates of oxygen uptake and of swelling in iso-osmotic solutions of ammonium salts, it is concluded that fat-cell mitochondria are permeable to the monocarboxylic acids, pyruvate and acetate; that in the presence of phosphate they are permeable to malate and succinate and to a lesser extent oxaloacetate but not fumarate; and that in the presence of both malate and phosphate they are permeable to citrate, isocitrate and 2-oxoglutarate. In addition, isolated fat-cell mitochondria have been found to oxidize acetyl l-carnitine and, slowly, l-glycerol 3-phosphate. 3. It is concluded that the major means of transport of acetyl units into the cytoplasm for fatty acid synthesis is as citrate. Extensive transport as glutamate, 2-oxoglutarate and isocitrate, as acetate and as acetyl l-carnitine appears to be ruled out by the low activities of mitochondrial aconitate hydratase, mitochondrial acetyl-CoA hydrolyase and carnitine acetyltransferase respectively. Pathways whereby oxaloacetate generated in the cytoplasm during fatty acid synthesis by ATP-citrate lyase may be returned to mitochondria for further citrate synthesis are discussed. 4. It is also concluded that fat-cells contain pathways that will allow the excess of reducing power formed in the cytoplasm when adipose tissue is incubated in glucose and insulin to be transferred to mitochondria as l-glycerol 3-phosphate or malate. When adipose tissue is incubated in pyruvate alone, reducing power for fatty acid, l-glycerol 3-phosphate and lactate formation may be transferred to the cytoplasm as citrate and malate.
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PMID:The intracellular localization of enzymes in white-adipose-tissue fat-cells and permeability properties of fat-cell mitochondria. Transfer of acetyl units and reducing power between mitochondria and cytoplasm. 439 82

Three collagenases were purified from the culture medium of human skin fibroblasts using ammonium sulfate fractionation, ion-exchange chromatography and gel filtration. The cationic collagenase had a molecular weight of 42,000; two anionic collagenases had molecular weights of 63,000 and 115,000. Preincubation of the individual collagenases with purified human and bovine platelet heparin binding proteins resulted in the inhibition of the two anionic activities, but only by bovine low heparin affinity platelet protein (beta-TG). Such inhibition was dose-dependent at the microgram level, was not antagonized by heparin, and persisted even when the collagenases had been transformed into their 53,000 and 105,000 forms through treatment with p-aminophenylmercuric acetate. Neither human nor bovine high heparin affinity platelet factors (PF-4) nor human low heparin affinity platelet protein (beta-TG) were inhibitory to any of the three collagenases studied. This suggests that the ability of platelet proteins to inhibit collagenase is specifically influenced by the ionic nature of the enzyme and this inhibition is specifically dependent upon the species and type of platelet protein.
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PMID:The interaction of platelet proteins with three fibroblast-derived collagenases. 608 70

A rat osteosarcoma cell clone (ROS 17/2), and osteoblast-enriched populations from rat calvaria cultured in the presence of concanavalin A, have been shown to produce latent collagenase and collagenase inhibitors. The enzymes and inhibitor activities from the ROS 17/2 cells were concentrated by ammonium sulphate precipitation and separated by gel filtration on AcA 54 resin. The size of the latent collagenase (Mr approximately equal to 58000) was reduced on conversion to active enzyme (Mr approximately equal to 48000) by p-aminophenylmercuric acetate. Latent and active forms of gelatinase activity, similar in size to the corresponding forms of collagenase, were also resolved. The collagenase inhibitor activity, which was sensitive to organomercurials, was recovered in two peaks (Mr approximately equal to 68000 and 30000). The active collagenase cleaved interstitial collagens (type I = III greater than II) producing typical 3/4 and 1/4 fragments. This activity was inhibited by the metal ion chelators ethylenediaminetetraacetic acid and o-phenanthroline. Additional specific cleavages of native collagen were also observed which, from the susceptibility of this activity to phenylmethylsulphonyl fluoride, leupeptin and antipain, suggested the presence of a second collagenolytic enzyme. This synthesis of collagenolytic enzymes by these osteoblast-like cells suggests that individual osteoblasts, like fibroblasts, are capable of both synthesizing and degrading their respective organic matrices in vivo.
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PMID:Synthesis of collagenase and collagenase inhibitors by osteoblast-like cells in culture. 609 78

The effects of human alpha 2macroglobulin (alpha 2M) on locomotion of human neutrophils and monocytes in micropore filter assays in vitro were studied. Native alpha 2M had no effect on cell locomotion. In contrast, alpha 2M which had been modified by interaction with proteases (trypsin, collagenase), or with ammonium sulphate, stimulated locomotion of both cell types. The degree of locomotory response induced in the cells by alpha 2M correlated closely with changes in molecular conformation of alpha 2M as estimated by measurements of binding of the fluorescent probe, 1-anilinonaphthalene-8-sulphonic acid (ANS). However neutrophil locomotion stimulated by modified alpha 2M appeared to be solely chemokinetic, whereas monocytes showed both chemokinetic and chemotactic responses to modified alpha 2M. The reason for this difference in response between the two cell types is unclear, but it is reflected in the lack of specific binding of alpha 2M-protease complexes by neutrophils. Earlier workers had shown specific binding of such complexes by mononuclear phagocytes.
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PMID:Effect of modified alpha 2macroglobulin on leucocyte locomotion and chemotaxis. 619 5

Two latent collagenases whose apparent molecular weights were ca. 150 000 and 60 000 have been detected in human leukocytes and the partial purification of the high molecular weight component was accomplished by the following steps: acetone precipitation, ammonium sulphate fractionation, gel filtration on Sephacryl S-200, chromatography on DEAE-Sepharose, and a final gel filtration on Sephacryl S-200. After activation by an activator extracted from human rheumatoid synovial fluid, the enzyme was able to cleave collagen into the classical 1/4 and 3/4 fragments, and was inhibited by chelating agents and other typical collagenase inhibitors.
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PMID:Partial purification and characterization of the high molecular weight latent collagenase from human leukocytes. 624 73

Glycoproteins from bovine aorta intima were isolated by a sequential digestion of the tissue with collagenase and elastase after extration of the tissue with saline. The proteins in the extracts were precipitated by (NH4)2SO4 and fractionated by Con-A sepharose affinity chromatography. The fractions were analyzed for their carbohydrate composition and by polyacrylamide disc gel electrophoresis. These studies show that considerable heterogeneity of aorta glycoproteins exists. Some of the glycoprotein materials are likely intimately associated with fibrous structures, collagen and elastin, of the aorta intima.
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PMID:Studies of glycoproteins from bovine aorta. 625 71

1. A latent collagenase (EC 3.4.24.3) has been isolated from rheumatoid synovial fluids and purified by (NH4)2SO4 precipitation and column chromatography, utilising Sephadex G-150, DEAE Sephadex A-50 and Sephadex G-100 superfine grade. 2. The final preparation activated by trypsin (EC 3.4.21.4) had a specific activity against thermally reconstituted collagen fibrils of 259 micrograms collagen degraded/min per mg enzyme protein, representing a nearly 800-fold increase over that of the original rheumatoid synovial fluid. 3. The latent collagenase preparation can be activated by trypsin and to some extent by HgCl2 but not by 3 M NaSCN, 3.5 M NaCl, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) or p-chloromercuribenzoate. 4. Inhibition studies and the acrylamide gel electrophoretic pattern of collagen degradation products showed that the trypsin-activated enzyme has the essential features of a neutral collagenase. 5. The molecular weights, determined by calibrated gel filtration, were 52 000 and 43 000 for the latent and the activated enzyme, respectively. 6. The nature of the latency of synovial fluid collagenase is discussed.
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PMID:A latent collagenase from rheumatoid synovial fluid. Purification and partial characterization. 625 72

A neutral protease has been extracted from the media of cultured metastatic tumor cells and purified approximately 1000 times after sequential ammonium sulfate fractionization, concanavalin A column chromatography, and molecular sieve chromatography. The protease has an apparent molecular weight of 70 000--80 000, is inactive at acid pH, requires trypsin activation, and is inhibited by ethylene-diaminetetraacetic acid but not by phenylmethanesulfonyl fluoride, N-ethylmaleimide, or soybean trypsin inhibitor. The enzyme produces specific cleavage products for both chains of pro type IV collagen isolated without pepsinization and apparently cleaves at one point in a major pepsin-extracted chain of placenta type IV collagen. The partially purified enzyme fails to significantly degrade other collagens or fibronectin under digestion conditions in which specific reaction products are produced for type IV collagen. The existence of this enzyme is significant since previously described animal collagenases fail to degrade type IV collagen. Such a type IV specific collagenase could play a role in tumor invasion and may be secreted by other cells such as endothelial cells, epithelial cells, and immune cells.
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PMID:Partial purification and characterization of a neutral protease which cleaves type IV collagen. 625 30


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