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)

Native type III collagen and procollagen were prepared from fetal bovine skin. Examination of the cleavage products produced by digestion with tadpole collagenase demonstrated that the three palpha1(III) chains of type III procollagen were linked together by disulfide bonds occurring at both the amino-terminal and carboxy-terminal portions of the molecule. Type III collagen contained interchain disulfide bonds only in the carboxy-terminal region of the molecule. After digestion of procollagen with bacterial collagenase an amino-terminal, triple-stranded peptide fragment was isolated. The reduced and alkylated chain constituents of this fragment had molecular weights of about 21 000. After digestion of procollagen with cyanogen bromide a related triple-stranded fragment was isolated. The chains of the cyanogen bromide fragment had a molecular weight of about 27 000. When the collagenase-derived peptide was fully reduced and alkylated, it became susceptible to further digestion with bacterial collagenase. This treatment released a fragment of about 97 amino acid residues which contained 12 cystein residues and had an amino acid composition typical for globular proteins. A second, non-helical fragment of about 48 amino acid residues contained three cysteines. This latter fragment is formed from sequences that overlap the amino-terminal region in the collagen alpha1(III) chain by 20 amino acids and possesses an antigenic determinant specific for the alpha1(III) chain. The collagenase-sensitive region exposed by reduction comprised about 33 amino acid residues. It was recovered as a mixture of small peptides. These results indicate that the amino-terminal region of type III procollagen has the same type of structure as the homologous region of type I procollagen. It consists of a globular, a collagen-like and a non-helical domain. Interchain disulfide bonding and the occurrence of cysteines in the non-helical domain are, however, unique for type III procollagen.
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PMID:Characterization of the amino-terminal segment in type III procollagen. 18 41

Endothelial cells isolated from bovine aorta synthesize and secrete type III procollagen in culture. The procollagen, which represents the major collagenous protein in culture medium, was specifically precipitated by antibodies to bovine type III procollagen and was purified by diethyl-aminoethylcellulose chromatography. Unequivocal identification of the pepsin-treated collagen was made by direct comparison with type III collagen isolated by pepsin digestion of bovine skin, utilizing peptide cleavage patterns generated by vertebrate collagenase, CNBr, and mast cell protease. The type III collagen was hydroxylated to a high degree, having a hydroxyproline/proline ratio of 1.5:1.0. Pulse-chase studies indicated that the procollagen was not processed to procollagen intermediates or to collagen. Pepsin treatment of cell layers, followed by salt fractionation at acidic and neutral pH, produced several components which were sensitive to bacterial collagenase and which comigrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with alpha A, alpha B, and type IV collagen chains purified from human placenta by similar techniques. Bovine aortic endothelial cells also secreted fibronectin and a bacterial collagenase-insensitive glycoprotein which, after reduction, had a molecular weight of 135,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (using procollagen molecular weight standards) and which was not precipitable by antibodies to cold-insoluble globulin or to alpha 2-macroglobulin. Collagen biosynthesis by these cells provides an interesting model system for studying the polarity of protein secretion and the attachment of cells to an extracellular matrix. The presence of type III collagen in the subendothelium and the specific interaction of this protein with fibronectin and platelets suggest the involvement of this collagen in thrombus formation following endothelial cell injury.
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PMID:Collagen synthesis by bovine aortic endothelial cells in culture. 39 Dec 67

Peptides corresponding to selected sequences of the alpha 1 chain of the COOH propeptide of type I and type III human procollagen were synthesized and used as antigens to develop polyclonal and monoclonal antibodies. The antibodies were shown to be epitope specific using a peptide-based solid phase enzyme-linked immunoadsorbent assay. The antibodies were specific for the appropriate procollagens and the COOH propeptides isolated from serum-free culture supernatants of human skin fibroblasts. The rabbit antisera directed to the type I synthetic peptide bound the intact procollagen molecule and both the procollagen alpha 1(I) and alpha 2(I) chains after the reduction of the disulfide bonds. In addition, the antisera bound intact type I COOH propeptide, generated by bacterial collagenase treatment of procollagen, and the individual chains of the propeptide after reduction. In contrast, a monoclonal antibody to the type I peptide was able to bind only to the reduced form of the COOH propeptide. Both rabbit polyclonal and murine monoclonal antibodies directed to the type III synthetic peptide bound the intact and the individual chains of type III procollagen as well as the intact and reduced forms of the type III COOH propeptide. The antibodies have been used to detect procollagen synthesis in two human osteosarcoma cell lines and the differential expression of procollagen in the culture medium of rat lung fibroblasts grown in the presence or absence of glucocorticoids.
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PMID:Detection of procollagen biosynthesis using peptide-specific antibodies. 169 17

When subconfluent cultures of primary human skin fibroblasts are incubated for 20 h in the presence of 5% neutral dextran, the newly synthesized procollagens are shifted from the medium into the pericellular matrix fraction. This is accompanied by an overall decrease by 30-50% in the secretion rates of these proteins, as indicated by the incorporation of tritiated proline into collagenase-sensitive macromolecules and by radioimmunoassays for the propeptide regions of type I and III procollagens. Inhibition of tyrosine sulphation in newly formed type III procollagen by NaClO3 does not change the distribution of this protein between the medium and matrix fractions either in the presence or in the absence of the polymer. Processing of type III procollagen at its N-terminus is incomplete in this situation, irrespective of whether the protein is present mainly in the medium or in the pericellular matrix. The concentrations of the mRNAs for the pro alpha 1(I)- and pro alpha 1(III)-chains of procollagens and for actin were monitored for up to 20 h; in the dextran-treated cultures these are not different from the corresponding concentrations in control cultures, indicating that the rapid down-regulation of the synthesis of type I and III procollagens in response to the enhanced pericellular matrix deposition induced by dextran is not due to transcriptional mechanisms.
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PMID:Effect of dextran on synthesis, secretion and deposition of type III procollagen in cultured human fibroblasts. 171 64

In addition to fibrosis in response to necrosis and inflammation, alcohol may promote fibrogenesis directly, resulting in pericellular, perisinusoidal and perivenular fibrosis, in association with increased collagen mRNA. Acetaldehyde (produced in increased amounts because of the selective induction of cytochrome P450IIE1) stimulates collagen formation from either myofibroblasts, Ito cells or fibroblasts. One postulated mechanism is adduct formation of acetaldehyde with intracellular proteins, possibly stabilized by the NADH generated upon ethanol oxidation. The latter also increases lactate which might inhibit proline oxidase activity and increase available proline. During the initial stage of alcohol consumption, collagenase activity is increased, in keeping with enhanced collagen production. In later stages, however, there is a secondary decrease of collagenase activity; its deficiency relative to synthesis may also promote collagen deposition. In the early stage of alcohol induced fibrosis, collagens type I and type III accumulate, whereas later, type I predominates. Both can be formed in vitro by Ito cells, but cultured hepatocytes produce mainly type III. Immunohistochemical techniques revealed the deposition of type III procollagen in the extrahepatic collagen fibrils. Its degradation, as well as its enhanced synthesis, contribute to the appearance of procollagen III peptides in the blood. Their measurement by Fab fragments of the antibody is useful to assess the degree of fibrosis and to detect perivenular fibrosis, a precirrhotic lesion. In the baboon model, polyunsaturated lecithin was found to be effective in opposing alcohol-induced fibrosis.
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PMID:Alcohol and fibrogenesis. 184 59

The effects of recombinant human Interleukin-1 alpha (IL-1 alpha), Interleukin-1 beta (IL-1 beta), and Tumor Necrosis Factor-alpha (TNF-alpha) on collagen biosynthesis were studied in vitro using dermal fibroblast cultures. Both forms of IL-1 and TNF-alpha induced a dose-dependent inhibition of both types I and III collagen synthesis, as measured by radioimmunoassay, gel electrophoresis, or collagenase-sensitive material. This effect was accompanied by a significant release of prostaglandin E2 into the culture medium. However, indomethacin, a potent inhibitor of prostaglandin synthesis, could not prevent the inhibitory effect of the three cytokines on collagen synthesis. Measurement of type I and type III procollagen mRNA levels in IL-1 treated cells revealed that both IL-1 alpha and IL-1 beta were potent enhancers of procollagen gene expression at pretranslational level. On the other hand, TNF-alpha was found to reduce the steady-state levels of type I and III procollagen mRNA in a dose-dependent manner. Quantitation of IL-1 beta and TNF-alpha transcripts following TNF-alpha treatment of fibroblasts indicated that this cytokine can induce IL-1 beta gene expression in these cells. By contrast, TNF-alpha mRNA remained at a constant level after TNF-alpha exposure. These data suggest that IL-1 and TNF-alpha, two cytokines that share several biologic activities, modulate collagen deposition in dermal fibroblasts by mechanisms that are clearly different: TNF-alpha appears to act at a transcriptional level to inhibit collagen synthesis, whereas IL-1 inhibitory action involves important translational regulation, still unknown, that counterbalances its stimulatory effect on procollagen mRNA levels. Moreover, our data suggest the existence of local fibroblastic cytokine production that may be involved in the modulation of extracellular matrix deposition.
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PMID:Comparative effects of interleukin-1 and tumor necrosis factor-alpha on collagen production and corresponding procollagen mRNA levels in human dermal fibroblasts. 199 84

The dermis of a child with Ehlers-Danlos syndrome type IV (EDS-IV) contained about 11% of the normal amount of type III collagen and cultured dermal fibroblasts produced a reduced amount of type III procollagen which was secreted poorly. Type III collagen produced by these cells contained normal and abnormal alpha-chains and cyanogen bromide peptides. The site of the structural defect in the abnormal alpha 1 (III) chains was localized to the region of Met797, which is at the junction of the two carboxyl-terminal CB5 and CB9 cyanogen bromide peptides. Chemical cleavage of heteroduplexes formed between EDS-IV mRNA and a normal cDNA clone covering the CB5 and CB9 region showed that about 100 nucleotides were mismatched. Sequencing of amplified and cloned cDNA spanning the mutant region revealed a 108 nucleotide deletion corresponding to amino acid residues Gly775 to Lys810. The deleted nucleotide sequence corresponded to sequences that, by analogy to the organization of the type I collagen genes, should be precisely encoded by exon 41 of the COL3A1 gene. Sequencing of amplified genomic DNA, prepared using disimilar amounts of primers specific for exons 41 and 42, displayed a base substitution (G-to-A) in the highly conserved GT dinucleotide of the 5' splice site of intron 41. Normal sequences were also obtained from the normal allele. It is likely that the GT-to-AT transition at the splice donor site of intron 41 generated an abnormally spliced mRNA in which sequences of exon 40 and 42 were joined together with maintenance of the reading frame. The corresponding peptide deletion included the cyanogen bromide cleavage site Met797-Pro798 and the mammalian collagenase cleavage site at Gly781-Ile782. These losses account for the resistance of EDS-IV collagen to cyanogen bromide and mammalian collagenase digestion. Cultured fibroblasts produced normal homotrimer, mutant homotrimer, and mixed heterotrimer type III collagen molecules. The mutant homotrimer molecules were the major pepsin-resistant species and about 69% of the alpha 1(III) mRNA was in the mutant form.
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PMID:A base substitution at a splice site in the COL3A1 gene causes exon skipping and generates abnormal type III procollagen in a patient with Ehlers-Danlos syndrome type IV. 214 68

The reported prevalence of interstitial lung disease in patients with rheumatoid arthritis has varied from 10% to 50%, yet less than 5% of patients with arthritis develop severe fibrosing interstitial lung disease. This suggests that subclinical disease may not always presage progressive disease. Bronchoalveolar lavage fluid from patients with rheumatoid arthritis and either clinically evident interstitial lung disease or subclinical disease was examined for the presence of factors with a putative role in the development of interstitial fibrosis. Patients with subclinical disease were identified by prospective radiographic and lung function screening of 93 patients with rheumatoid arthritis. Fourteen patients were identified in this manner and an association between subclinical disease and smoking history was noted. Eleven patients with established interstitial lung disease had increased neutrophils (p less than 0.05), collagenase, and type III procollagen N terminal peptide levels (p less than 0.01) in the bronchoalveolar lavage fluid. Preliminary characterisation of the bronchoalveolar lavage collagenase suggested that it originated from neutrophils. Ten patients with subclinical interstitial lung disease underwent bronchoalveolar lavage. Of these, one had increased neutrophils and two had increased collagenase concentrations--abnormalities associated with advanced interstitial lung disease and a poor prognosis. These results suggest that in arthritis patients with evidence of subclinical pulmonary interstitial disease bronchoalveolar lavage might be useful in identifying those who may require careful monitoring in the hope that early treatment will prevent severe fibrosis.
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PMID:Bronchoalveolar lavage in patients with mild and severe rheumatoid lung disease. 216 54

Previous observations (Stolle, C.A., Pyeritz, R.E., Myers, J.C., and Prockop, D.J. (1985) J. Biol. Chem. 260, 1937-1944) indicated that fibroblasts from a proband with dominantly inherited Ehlers-Danlos syndrome type IV synthesized type III procollagen with a structural defect near the collagenase cleavage site at amino acid 781 and near the trypsin-sensitive site at 789. The type III procollagen was unusually sensitive to proteinases and cleaved by trypsin into a three-quarter fragment at 0 degrees C. Here we demonstrate that the mutation in the type III procollagen gene is a single base mutation that converts the codon for glycine at amino acid 790 of the alpha 1(III) chain to a codon for serine. The mutation probably makes the procollagen molecule unusually sensitive to proteases because it causes local unfolding of the triple helix and exposes the adjacent arginine residue. The results provide the first indication that not all glycine substitutions in the triple helices of fibrillar collagens are equivalent in terms of their effects of the biological function of the molecule.
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PMID:A single base mutation that substitutes serine for glycine 790 of the alpha 1 (III) chain of type III procollagen exposes an arginine and causes Ehlers-Danlos syndrome IV. 249 73

Confluent cultures of normal human skin fibroblasts were labelled overnight with [35S]sulphate, and the incorporation of the isotope into type III procollagen, secreted into the medium, was verified by radioimmunoassay and immunoprecipitation after removing the heavily sulphated proteoglycans by anion-exchange chromatography. Type III procollagen and its pro and pN alpha chains were visualized in fluorographs of the immunoprecipitates. The labelled procollagen could be isolated by a combination of ion-exchange chromatography and gel filtration and was found to contain tyrosine O-sulphate, which was identified by thin-layer electrophoresis after Ba(OH)2 hydrolysis. The regions sulphated in the type III procollagen molecule were susceptible to pepsin digestion. Digestion with purified bacterial collagenase at +37 degrees C produced a labelled fragment that was recognized by antibodies against the aminoterminal propeptide of type III procollagen, indicating that the sulphated tyrosine residues are located either in this propeptide or in the non-helical telopeptide region of the type III collagen molecule proper. Sulphation of tyrosine residues is a new post-translational modification in procollagen, which could be involved in the regulation of the processing of type III procollagen into collagen and thus affect the formation of collagen fibres.
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PMID:Incorporation of sulphate into type III procollagen by cultured human fibroblasts. Identification of tyrosine O-sulphate. 300 90


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