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)

Thioglycolate-stimulated mouse peritoneal macrophages secrete a Proteinase which degrades insoluble elastin. There is little elastase activity in cell lysates but the bulk of the enzyme accumulates extracellularly during culture in serum-free medium. The secretion of elastase is sustained for over 12 days in culture and continued secretion of elastase requires protein synthesis. Unstimulated macrophages secrete very little elastase activity but can be triggered to secrete higher levels of this enzyme by phagocytosis and intracellular storage of latex particles. The macrophages elastase is a distinctive proteinase differing from the elastases of pancreas and granulocytes and is distinct from the other secreted proteinases of macrophages, namely, collagenase and plasminogen activator. The macrophages elastase is a serine proteinase and is inhibited by di-isopropyl phosphoro-fluoridate, ovoinhibitor, EDTA, dithiothretiol, and serum. Its activity is little affected by soybean trypsin inhibitor, turkey ovomucoid and chloromethyl ketones derived from tosyl lysine, tosyl phenylalanine, and acetyltetra alanine. Hydrolysis by macrophage elastase of chromogenic ester substrates for pancreatic elastase could not be detected. Elastase secretion by stimulated macrophages exceeds that by primary and established fibroblast cell strains. It is likely that elastase secretion by macrophages plays a major role in the pathogenesis of chronic destructive pulmonary diseases such as emphysema.
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PMID:Elastase secretion by stimulated macrophages. Characterization and regulation. 16 96

Sequencing of chymotrypsin, trypsin, collagenase- and hydroxylamine-derived peptides, using the automated Edman degradation procedure, yielded the complete amino acid sequence of alpha2-CB4 from calf skin collagen (321 residues). Together with the data from earlier work, an uninterrupted sequence in the helical region of the alpha2-chain from residues 1-393 is now known. Glycine is found in every third position of the peptide. Hydroxylation of proline and lysine occurs only in the Y-position of the triplet Gly-X-Y and is not complete in every position. Some residues, such as glutamic acid, leucine, phenylalanine and arginine, are distributed non-randomly between the X and Y-positions and this non-random distribution is different in the alpha1 and alpha2-chains. Comparison of the N-terminal 393 residues from the helical region of the alpha1 and alpha2-chains revealed a nearly identical distribution of charged polar residues arginine, lysine, glutamic and aspartic acids. The distribution of the triplet Gly-Pro-Hyp is simialr in both chains. The remaining residues in the alpha2-chain exhibit a high degree of substitutions when compared with those in the alpha1-chain. Approximately one in every two residues in both the X and Y-positions are substituted.
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PMID:The covalent structure of collagen. The amino-acid sequence of alpha2-CB4 from calf-skin collagen. 17 31

Techniques are descirbed for the isolation and culture of endothelial cells from the lungs of small animals. The cells are collected by retrograde perfusion of blood-free lungs with buffered saline containing collagenase. The cells are characterized by light microscopy, electron microscopy of thin sections and surface replicas, and by the presence of angiotensin-converting enzyme (ACE). ACE was assayed using 3H-benzoyl-Phe-Ala-Pro as substrate and was localized by indirect immunofluorescence using guinea pig endothelial cells incubated with rabbit antibodies to guinea pig lung ACE followed by goat anti-rabbit globulins conjugated to fluorescein. Thus, endothelial cultures can be established using small animals commonly employed in studies of pulmonary processing of vasoactive substances.
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PMID:Isolation and culture of endothelial cells from the lungs of small animals. 22 81

Pancreatic islets of Wistar rats were isolated by collagenase digestion and incubated with [3H]-L-phenylalanine. Using a specific somatostatin antiserum radioactivity was found in the antibody-antigen-complex. The radioactivity was displaced by unlabelled somatostatin. These findings give the first evidence for the biosynthesis of somatostatin or somatostatin-like peptides in mammalian pancreatic islets.
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PMID:Biosynthesis of somatostatin in pancreatic islets of Wistar rats. 37 44

Converting enzyme activity was studied in endothelial cells, isolated from pig pulmonary arteries and aorta by exposure to collagenase. The measure was based on the release of His-Leu from Z-Phe-His-Leu was related to the DNA content of the cellular suspension. The same activity was found in the two types of endothelium: 1 nmol His-Leu/mug DNA per 30 min. Subendothelial cells showed a very low activity, amounting to 10% of the value found for the endothelium. The enzyme activity was 2inhibited by the nonapeptide SQ 20881, EDTA, and the lack of Cl- in the same fashion for the two types of endothelium. The presence of another enzyme hydrolyzing His-Leu was detected in both endothelial populations. Isolated fragments of plasma membrane, however, exhibited only converting enzyme activity. It can be concluded that endothelial cells isolated from large vessels of the pulmonary and the systemic circulations have similar properties when dipeptidyl carboxypeptidase activity is measured.
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PMID:Converting enzyme activity in endothelial cells isolated from pig pulmonary artery and aorta. 40 22

1. Isolated parenchymal cells were prepared by collagenase perfusion of livers from fed rats that had been previously injected with [(3)H]leucine to label liver proteins. When these cells were incubated in a salts medium containing glucose, gelatin and EDTA, cellular integrity was maintained over a period of 6h. 2. Cells incubated in the presence of 2mm-leucine to minimize radioactive isotope reincorporation released [(3)H]leucine into the medium at a rate accounting for the degradation of 4.5% of the labelled cell protein per h. 3. Degradation of [(3)H]protein in these cells was inhibited by insulin and by certain amino acids, of which tryptophan and phenylalanine were the most effective. 4. Protein degradation was decreased by several proteinase inhibitors, particularly those that are known to inhibit lysosomal cathepsin B, and by inhibitors of cell-energy production. 5. Ammonia inhibited degradation, but only at concentrations above 1.8mm. Aliphatic analogues of ammonia were effective at lower concentrations than was ammonia. 6. High concentrations of ammonia inhibited degradation by 50%. The extent of this inhibition could not be increased further by the addition of the cathepsin B inhibitor leupeptin, which by itself inhibited degradation by approx. 30%. 7. The sensitivity of proteolysis in isolated hepatocytes to these various inhibitory agents is discussed in relation to their possible modes of action.
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PMID:Inhibition of protein degradation in isolated rat hepatocytes. 88 Feb 45

Porcine type-I collagenase (Colg-1) was produced as a fusion protein in Escherichia coli using the pAX5 expression vector. The fusion protein consists of beta-galactosidase at the N terminus joined to a collagen hinge region and a blood-coagulation factor Xa cleavage site linked to Colg-1. Recombinant collagenase (reColg-1) was biologically active in the form of a fusion protein and could be released by treatment with factor Xa to yield Colg-1 with the authentic N terminus (phenylalanine) found in vivo. The results show that reColg-1 produced in E. coli is folded correctly, cleaves type-I collagen into 1/4 and 3/4 fragments at the characteristic Colg-sensitive site, and is produced at high enough levels to generate a source of recombinant enzyme for x-ray crystallography studies.
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PMID:Production in Escherichia coli of porcine type-I collagenase as a fusion protein with beta-galactosidase. 131 1

Rat calvaria bone cells isolated by collagenase digestion form a bone-like matrix which mineralizes in vitro in the presence of beta-glycerophosphate, in less than 2 weeks. The purpose of this work was to investigate, in this mineralizing rat osteoblastic cell culture, the synthesis of collagen, osteocalcin, and bone alkaline phosphatase (ALP). The results obtained indicate (1) After 15 days in culture, the extracellular-matrix contains collagen type I, V, and to some extent type III. Metabolic labeling at day 14, during the phase of nodules mineralization as well as new nodules formation, shows that collagen types I and type V are synthesized; (2) During the phase of cell growth, no osteocalcin could be detected in the medium, however, at the point of nodule formation, the osteocalcin level reached values of 3.55 +/- 1.39 ng/ml, followed by a 30-fold increase after nodules became mineralized. At day 14, after metabolic labeling, de novo synthesized osteocalcin was chromatographed on an immunoadsorbing column. With urea-SDS PAGE the apparent molecular weight was determined to be 9,000 daltons. (3) Specific activity of ALP was found to be 10 nmol/min/mg of proteins at cell confluence. At day 15, when nodules are mineralized, this activity was increased by 40-fold. The Michaelis constant was 1.58 10(-3) M/L. ALP was inhibited by L-homoarginine and levamisole but not by L-phenylalanine. ALP was shown to be heat sensitive at 56 degrees C with two slopes of inhibition.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Expression of collagen, osteocalcin, and bone alkaline phosphatase in a mineralizing rat osteoblastic cell culture. 137 88

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

Our investigations indicate that a variety of neutral serine proteases exist in highly purified, IL-2-activated rat NK (A-NK) cells. These enzymatic activities are not restricted to only cytolysin-containing granules and are not defined by only the assay of N-alpha-benzyloxycarbonyl-L-lysine thiobenzylesterase activity. These activities, which we term A-NKP 1, A-NKP 2, A-NKP 3, and A-NKP 4, cleave, respectively, the following fluorogenic peptide substrates: Boc-Phe-Ser-Arg-7-amino-4-methylcoumarin (AMC, trypsin-like); Suc-Ala-Ala-Phe AMC (chymotrypsin-like); Suc-Gly-Pro-Leu-Gly-Pro AMC (collagenase-like), and Z-Phe-Arg AMC (another trypsin-like enzyme). The proteases A-NKP 1, A-NKP 2, and A-NKP 3 are not cell surface-associated and appear to be cytosolic as defined by isopycnic sucrose density gradient centrifugation. In contrast, A-NKP 4 appears to be located in lysosomes. Treatment of rat A-NK cells with protease inhibitors that inhibit A-NKP 2 and A-NKP 3 also substantially inhibit A-NK cell-mediated cytotoxicity against both NK-sensitive and -resistant targets (YAC-1 and P815, respectively). These results indicate that A-NKP2 and A-NKP 3 may play a role in IL-2-activated NK cell-mediated cytotoxicity. A variety of proteolytic enzymes, in addition to granzymes, therefore exist in A-NK cells. Our studies indicate that a prerequisite to a thorough understanding of the role of proteases in killer cell function is the investigation of several classes of enzymes in addition to granzymes contained in lytic granules.
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PMID:Nongranular proteolytic enzymes of rat IL-2-activated natural killer cells. I. Subcellular localization and functional role. 151 70


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