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 processing of type III and type I procollagen molecules in cultured bovine aortic smooth muscle cells was investigated. The molecular identities of the processing intermediates of type III and type I procollagen were characterized by analysis of the radioactive collagenous components using mammalian collagenase and pepsin digestions and cyanogen bromide peptide mapping. The results indicate that the processed intermediates for procollagen type III and type I are their respective pC components. Although the processing pathways for both collagen types are the same, data from pulse-chase experiments suggest that the rates at which the processing occurs are different. Type I procollagen is processed more rapidly to its intermediate than is type III procollagen. The type I pC intermediate is almost completely processed to alpha-chains and a significant portion of these fully processed molecules remains in a soluble form even after 11 h. In the same time period, the type III pC intermediate is slowly converted to alpha-chains. Since beta-aminopropionitrile was not employed in these studies, significant accumulation of collagen chains into the insoluble extracellular matrix was observed during the chase period.
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PMID:Processing of procollagen types III and I in cultured bovine smooth muscle cells. 674 44

Activation of bovine plasma prekallikrein was investigated with several proteinases. Highly purified bovine plasma prekallikrein was rapidly activated to kallikrein [EC 3.4.21.8] by bovine activated Hageman factor, trypsin [EC 3.4.21.4] and Pronase P (proteinases from Streptomyces griseus) and more gradually by papain [EC 3.4.22.2] and ficin [EC 3.4.22.3]. Activation of prekallikrein was also observed with bovine plasmin [EC 3.4.21.7], but not with bovine clotting factors Xa (Stuart factor) [EC 3.4.21.6] and IXa (Christmas factor) or thrombin [EC 3.4.21.5]. Urokinase [EC 3.4.99.26], Reptilase, collagenase [EC 3.4.24.3], elastase [EC 3.4.21.11], alpha-chymotrypsin [EC 3.4.21.1], Nagarse [EC 3.4.21.14], and stem bromelain [EC 3.4.22 4] did not convert prekallikrein to kallikrein. Plasma kallikrein activated to Hageman factor released kinin rapidly from bovine high molecular weight (HMW) kininogen. However, from bovine low molecular weight (LMW) kininogen, liberation of kinin was extremely slow. The kallikrein activity was inhibited by soybean trypsin inhibitor (SBTI), Trasylol, diisopropylfluorophosphate (DFP), and N-alpha-tosyl-L-lysine chloromethylketone (TLCK), but not by egg-white trypsin inhibitor (EWTI), lima bean trypsin inhibitor (LBTI), heparin or hexadimethrine bromide (Polybrene). The kallikrein formed an enzyme-inhibitor complex with SBTI and Trasylol, but not with LBTI. Prekallikrein did not react with SBTI. Prekallikrein consists of a single polypeptide chain of molecular weight about 90,000, as estimated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Activation of prekallikrein by Hageman factor was found to involve cleavage of the single peptide bond on the disulfide-bridged polypeptide chain, and no change of molecular weight was observed during the activation. The peptide bond cleaved in prekallikrein by the activation was an Arg-X peptide bond on a disulfide-bridged polypeptide chain.
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PMID:Studies on prekallikrein of bovine plasma. II. Activation of prekallikrein with proteinases and properties of kallikrein activated by bovine Hageman factor. 676 24

Dermal fibroblasts in culture from a woman with a mild to moderate form of osteogenesis imperfecta synthesize two species of the pro alpha 2-chain of type I procollagen. One chain is normal. The abnormal chain has a slightly faster mobility than normal during electrophoresis in sodium dodecyl sulfate polyacrylamide gels. Analysis of cyanogen bromide peptides of the pro alpha-chain, the alpha-chain, and of the mammalian collagenase cleavage products of the pro alpha- and alpha-chains indicates that the abnormality is confined to the alpha 2(I)CB4 fragment and is consistent with loss of a short triple-helical segment. Type I collagen production was decreased, perhaps because the molecules that contained the abnormal chain were unstable, with a resultant alteration in the ratio of type III to type I collagen secreted into culture medium. Collagen fibrils in bone and skin had a normal periodicity but their diameters were 50% of control; the bone matrix was undermineralized. The structural abnormality in the alpha 2(I)-chain in this patient may affect molecular stability, intermolecular interactions, and collagen-mineral relationships that act to decrease the collagen content of tissues and affect the mineralization of bone.
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PMID:Abnormal alpha 2-chain in type I collagen from a patient with a form of osteogenesis imperfecta. 682 30

Cells in culture from a woman with a variety of the Marfan syndrome produce two species of the alpha 2 chains of type I collagen. One alpha 2 chain appears normal; the abnormal chain has a higher apparent molecular weight than normal and migrates more slowly during electrophoresis in sodium dodecyl sulfate/polyacrylamide gels. A similar change in electrophoretic behavior is seen in the prepro alpha 2 chain and the pN alpha 2 chain (which contains the amino-terminal extension). Asymmetric cleavage of the pepsin-treated procollagens with a fibroblast collagenase locates the abnormal segment amino terminal to the cleavage site, and analysis of cyanogen bromide peptides of collagenase cleavage peptides and of whole collagens indicates that the abnormal segment is in either the alpha 2CB3 peptide or the short segment of alpha 2CB5 amino terminal to the collagenase site of the altered alpha 2 chain. The higher apparent molecular weight is consistent with the insertion of a small peptide fragment of approximately 20 amino acids. This alteration in chain size has marked effects on crosslinking because collagen from the patient's skin was 5-10 times more extractable in nondenaturing solvents than that from control skins. Although the abnormal chain was not found in several other individuals with the Marfan syndrome, these findings suggest that the phenotype may be the expression of a variety of primary structure alterations in the chains of type I collagen that interfere with normal crosslink formation.
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PMID:Marfan syndrome: abnormal alpha 2 chain in type I collagen. 695 Apr 13

DNA synthesis in rat hepatocytes, from livers regenerating after 70% hepatectomy, was assessed by flow cytometric determination of nuclear DNA content and by incorporation of [3H]thymidine. Parenchymal liver cells were isolated by collagenase perfusion and low-speed centrifugation. Nuclei from the isolated cells were prepared for flow cytometry by a treatment with detergent, pepsin and RNase, and stained with ethidium bromide. Parallel samples of cells were incubated with [3H]thymidine and analysed for rate of incorporation of radioactivity into DNA and for labelling index determination. The flow cytometric measure of the replicative response, i.e. the presence of cells with S-phase DNA content within the diploid and tetraploid cell populations, was compared with the incorporation of [3H]thymidine. For each of fourteen animals, including two control rats and twelve partially hepatectomized animals killed either before (at 13 hr after hepatectomy), at the onset (16 and 18 hr) or at the peak (24 hr) of regenerating activity, a fairly good correlation was found between the different methods. Satisfactory resolution of the flow cytometric detection of S-phase cells was indicated by a sorting experiment using an Ortho (system 50-H) cell sorter which demonstrated that after [3H]thymidine injection in vivo 88% of the diploid and 84% of the tetraploid S-phase nuclei were labelled, while labelling in the G1-fractions was only 2 and 7%, respectively.
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PMID:The proliferation response of rat liver parenchymal cells after partial hepatectomy. A methodological study comparing flow cytometry of nuclear DNA content and in vivo and in vitro uptake of thymidine. 712 1

Distribution of nuclear ploidy in female mouse hepatocytes was measured cytofluorimetrically using ethidium-bromide-stained hepatocyte nuclei liberated by in situ collagenase perfusion of the liver via the portal vein. After i.v. administration of lead acetate or an i.p. general anaesthetic (Valium and Hypnorm), rapid shifts of 8N and 4N nuclei to lower ploidy levels were recorded. It was possible to block the ploidy changes with i.p. colchicine, although no blocked metaphases were observed histologically. These observations were consistent with an earlier report on changes in nuclear ploidy induced by carbon tetrachloride (Steele et al., 1981b). It is speculated that these changes in ploidy may represent an early response of the hepatocytes to stimulation--an aspect of liver behaviour of which we were not previously aware.
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PMID:Some flow-cytofluorimetric studies of the nuclear ploidy of mouse hepatocytes: iii. further observations on early changes in nuclear ploidy of mouse hepatocytes following various experimental procedures. 715 May 7

The collagenase domain of bovine glomerular basement membrane was isolated in soluble form after limited digestion with pepsin. Gel filtration chromatography of the domain under denaturing conditions revealed that most of the polypeptide constituents exhibit apparent molecular weights greater than the type I collagen beta-chain, while approximately 15% are similar in size to that of alpha-chain. Carboxymethyl cellulose chromatography of the alpha-size region revealed that 70% of the protein was polypeptide XIV, as previously designated (West, T. W., Fox, J. W., Jodlowski, M., Freytag, J. W., and Hudson, B. G. (1980) J. Biol. Chem. 255, 10451-10459). This polypeptide exhibits an apparent molecular weight of 102,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. An absolute molecular weight value of 86,000 was determined by sedimentation equilibrium ultracentrifugation in 6 M guanidine hydrochloride. About 15% of the mass is carbohydrate which exists in the form of glucosylgalactosylhydroxylysine. Thus, the polypeptide backbone has a molecular weight of 73,000, a value which is considerably smaller than the alpha-chains of classical collagen. The amino acid and carbohydrate composition and cyanogen bromide patterns indicate that polypeptide XIV has a structure similar to that of C-chain or alpha 1 (IV) collagen which has been identified in other tissues. In addition, the cyanogen bromide pattern of the entire collagenous domain is similar to that of polypeptide XIV, suggesting that the latter is a structural segment of many of the higher molecular weight components.
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PMID:Bovine glomerular basement membrane. Characterization of an alpha-size collagenous polypeptide. 725 9

Mature mice have a large proportion of their hepatocyte nuclei in polyploid states (tetraploid and octaploid), and this is more prominent in females. We measured nuclear ploidy distribution cytometrically using ethidium bromide-stained hepatocyte nuclei liberated by in situ collagenase perfusion of the liver via the portal vein. After s.c. administration of 0.2 ml carbon tetrachloride the ploidy distributions of 8-month-old female mice changed from a control of 35% 2N, 45% 4N, and 20% 8N to 54% 2N, 45% 4N and 1% 8N at 6 h, and 65% 2N, 35% 4N and 0% 8N at 24 h. By 72 h 92% of the nuclei were diploid. These changes preceded any changes in mitotic index and S-phase index (3H-TdR autoradiographs). Histology confirmed the loss of higher-ploid nuclei but without mitotic figures or selective cell necrosis to account for the observations. Cleaved nuclei were prominent in sections of liver examined 3 h after CCl4 administration and suggested division of polypoid nuclei that had undergone prior segregation of chromatids and had presumably been arrested in telophase.
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PMID:Some flow cytofluorimetric studies of the nuclear ploidy of mouse hepatocytes. II. Early changes in nuclear ploidy of mouse hepatocytes following carbon tetrachloride administration: evidence for polyploid nuclei arrested in telophase. 729 42

Studies were undertaken to define the molecular size of the elastin primary gene product. Translation of chick aortic messenger ribonucleic acid (mRNA) in an mRNA-dependent reticulocyte lysate resulted in the synthesis of two major proteins of 70 000 and 73 000 molecular weights. Both proteins were shown to be soluble forms of elastin by isotope incorporation, immunoprecipitation, collagenase and cyanogen bromide sensitivity, and two-dimensional gel electrophoresis. The 70 000-dalton protein behaves similarly to authentic tropoeleastin in sodium dodecyl sulfate gel electrophoresis. There was no evidence for a high molecular weight form of soluble elastin, although procollagen chains were indirectly identified among the aortic mRNA-directed translation products. The same molecular size proteins were also seen in organ cultures of chick embryonic aortas labeled with [3H]valine. However, the 73 000-dalton protein was not extractable in a neutral salt buffer but was found only if the aortas were extracted with urea in the presence of reducing and alkylating reagents. The results from these studies suggest that elastin is first synthesized as two distinct polypeptide chains which differ slightly in size and overall charge. The possibility that these two proteins may associate posttranslationally to form a dimer prior to secretion is postulated to explain the existence of a putative proelastin molecule seen in other systems.
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PMID:Translation of chick aortic elastin messenger ribonucleic acid. Comparison to elastin synthesis in chick aorta organ culture. 735 64

Human skin fibroblasts in culture have previously been shown to synthesize genetically distinct procollagens type I and type III. In the present study, cultured human skin fibroblasts were incubated under conditions optimized for synthesis of these procollagens in medium containing [3H]proline. The newly synthesized type I and type III 3H-labeled procollagens in the culture medium were then isolated as native proteins by DEAE-cellulose chromatography, or by gel filtration and SDS-polyacrylamide slab gel electrophoresis under denaturing conditons after limited pepsin proteolysis. The chromatographic procedures were optimized to yield reliable and reporducible results with good recoveries. The isolated procollagens were identified by cyanogen bromide peptide mapping and characterized by cleavage with highly purified collagenase synthesized by human skin fibroblasts. Assay of the relative synthesis of type I/III procollagens by normal human skin fibroblasts using DEAE-cellulose chromatography indicated that 80% of the procollagen in the medium was type I while the remaining 20% consisted of type III. When the ratio of newly-synthesized type I/III collagens was estimated by gel filtration or using SDS-polyacrylamide slab gel electrophoresis after limited pepsin proteolysis, relatively fewer type III collagen alpha-chains were recovered. This observation suggests that some of the type of the type III collagen molecules are in a conformation which is less resistant to digestion by pepsin than the triple-helix of type I procollagen. The coefficient of variation for the relative synthesis of type I and type III procollagens by control cultures was relatively small (16%), indicating that the phenotypic expression of type I and type III procollagen genes, under optimized culture conditions, is under a relatively tight control. The results further suggest that the optimized methodology developed for assay of the relative synthesis of type I and type III procollagens and collagens by cultured human skin fibroblasts can be utilized in studies on collagen aberrations in acquired and inherited diseases of connective tissue.
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PMID:Collagen biosynthesis by human skin fibroblasts. II. Isolation and further characterization of type I and type III procollagens synthesized in culture. 741 91


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