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Query: EC:5.99.1.2 (topoisomerase)
 9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Calf thymus DNA topoisomerase I, which belongs to the eukaryotic type I topoisomerases, is in a typical preparation purified as a set of five major polypeptides with Mr between 70000 and 100000. At least four of these proteins have binding affinity for DNA as was shown by incubating them with radioactive single-stranded DNA after separation in dodecylsulfate polyacrylamide gels and blotting onto nitrocellulose filters. That these polypeptides have DNA relaxing activity was directly demonstrated with protein extracted from single bands of dodecylsulfate/polyacrylamide gels. We consider the 100000-Mr protein to be the native enzyme. The smaller components are catalytically active fragments of the native topoisomerase most probably arising from limited proteolysis either within the nucleus or during the purification of the enzyme. In two-dimensional non-equilibrium pH-gradient electrophoresis gels the topoisomerase size variants exhibit apparent pI values between 8.1 and 8.3, with small but distinct differences between the components. The calf thymus topoisomerase I, upon binding to phage fd-DNA, protects a stretch of 15-25 nucleotides against digestion with DNase I.
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PMID:Characterisation of size variants of type I DNA topoisomerase isolated from calf thymus. 609 Jan 40

Chromatin-associated DNA topoisomerase I activity was measured in human diploid fibroblasts during in vitro aging. No difference was detected as a function of cell age in the nicking and the closing activities of the DNA-unwinding enzyme. The capacity of type-I topoisomerase to relax superhelical DNA molecules was, however, increased in aged cells. An age-related increase in nucleoprotein content was also observed.
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PMID:Increased DNA topoisomerase I activity in aging human cell chromatin. 609 22

Mutations in top, the structural gene for Escherichia coli DNA topoisomerase I, have been identified and mapped at 28 min on the chromosome, near cysB. Strains carrying deletions of the top gene are viable. The top mutations, however, do exert pleiotropic effects on transcription and transposition. Mutants lacking DNA topoisomerase I have a more rapid rate of induction and a higher level of catabolite-sensitive enzymes including tryptophanase and beta-galactosidase. This general activation of transcription by top mutations can be attributed to an increase in the negative superhelicity of the DNA in vivo when the topoisomerase activity is abolished. The frequency of transposition of Tn5, a transposon carrying kanamycin resistance, is decreased by a factor of 40 or more in top mutants. A direct or indirect role of the topoisomerase in transposition is discussed. The transposition frequency of Tn3, however, is not dependent on top. Based on the studies of the E. coli top mutants, it appears that the supX gene, which was originally studied in Salmonella typhimurium [Dubnau, E. & Margolin, P. (1972) Mol. Gen. Genet. 117, 91-112] is likely to be the structural gene for DNA topoisomerase I.
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PMID:Mutations in the gene coding for Escherichia coli DNA topoisomerase I affect transcription and transposition. 626 7

Activity of DNA topoisomerase I has been characterized in extracts of mitochondria purified from Xenopus laevis oocytes. Several lines of evidence have been obtained for the intramitochondrial localization of the enzyme. The mitochondria-associated of DNA topoisomerase I represents 1% of the activity recovered from a total ovary population of oocytes. The enzymes has been purified by DEAE-cellulose, phosphocellulose and double-stranded DNA cellulose chromatography and its properties compared to those of its nuclear-cytosolic counterpart purified by the same purification steps. Both enzymes appear to possess very similar if not identical physico-chemical and catalytic properties. Neither enzyme shows DNA gyrase activity and they are both equally sensitive towards trypanocide drugs such as ethidium bromide and berenil. The mitochondrial enzyme is able to remove positive superhelical turns and possibly acts as a swivelase during replication of mitochondrial DNA. These results confirm the pioneering work of Fairfield et al. (1979, J Biol. Chem. 254, 9354) on the presence of a DNA topoisomerase I in mitochondria. However our results support the conclusion that in Xenopus laevis oocytes the mitochondrial and nuclear cytosolic DNA topoisomerase I are in all likelihood one and the same enzyme.
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PMID:DNA topoisomerase I from mitochondria of Xenopus laevis oocytes. 626 55

The activity of DNA topoisomerase I(DNA nicking-closing enzyme) was analysed in cytoplasmic and nuclear extracts of six independently derived Fanconi and four normal fibroblast cell lines. In all experiments the total cellular activity was predominantly found in the nuclear extracts (88-100%). In addition, a minor proportion of the enzyme (up to 12%) was randomly present in some of the cytoplasmic fractions of both Fanconi and normal fibroblasts. These results indicate that Fanconi's anaemia is probably not due to or accompanied by a maldistribution of topoisomerase I between nuclei and cytoplasm.
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PMID:Intracellular distribution of DNA topoisomerase I in fibroblasts from patients with Fanconi's anaemia. 629 17

A DNA topoisomerase activity is found to be associated with the nucleosomes released by the Staphylococcal nuclease digestion of HeLa nuclei. Such an association is found to be salt dependent. A number of criteria have established that this DNA topoisomerase activity is due to HeLa topo I (Liu, L. F. and Miller, K. G. (1980) Proc. Natl. Acad. Sci. USA 78, 3489-3491). A similar association has been demonstrated from the in vitro studies using purified mononucleosomes and eukaryotic DNA topoisomerase I. Nonhistone HMG proteins and histone H1 are found to stimulate topoisomerase activity in vitro and form tight complexes with eukaryotic DNA topoisomerase I. The intimate interactions of topoisomerase I with chromosomal proteins and nucleosomes may be an essential feature of the topoisomerase function in vivo.
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PMID:Association of eukaryotic DNA topoisomerase I with nucleosomes and chromosomal proteins. 629 26

DNA winds about itself in a right-handed or left-handed fashion at several structural levels. The double helix is generally right-handed and is given a (+) sign by convention, whereas supercoiling of the helix axis is always (-) in the cell. The winding in higher -order forms such as knots and catenanes is unknown, and this has impeded elucidation of the mechanisms of their formation and resolution by replication, recombination and topoisomerase action. We introduce here a procedure for determining the handedness of DNA winding by inspection of electron micrographs of DNA molecules coated with Escherichia coli RecA protein. We demonstrate the validity of the method and show that DNA topoisomerase I of E. coli generates an equal mixture of (+) and (-) duplex DNA knots, and that one product of recombination by resolvase of transposon Tn3 (refs 8, 9) is a catenane of uniquely (+) sign.
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PMID:Determination of the absolute handedness of knots and catenanes of DNA. 630 70

We have previously shown that a DNA topoisomerase I from mouse mammary carcinoma cells is inhibited by heparin. Taking advantage of this enzyme-heparin interaction, we developed a rapid and efficient method of purification of this enzyme to near homogeneity by extraction of chromatin with 0.15 M phosphate buffer followed by two-step column chromatography on heparin-Sepharose and phenyl-Sepharose. Electrophoresis on sodium dodecyl sulfate-polyacrylamide gels revealed that the final preparation is composed of two polypeptides with apparent Mr approximately 98,000 (p98) and 102,000 (p102), p98 comprising 70% and p102 30%. Extraction and renaturation of the polypeptides from the gel shows that both p98 and p102 seem to possess topoisomerase activity. Partial proteolytic digestion of p98 and p102 with Staphylococcus aureus V8 and chymotrypsin yielded a series of identical peptides, indicating that the two polypeptides are structurally related. The enzyme sedimented through sucrose density gradient with s20,w of 4.0 S, and thus is monomeric in solution.
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PMID:Rapid purification and characterization of DNA topoisomerase I from cultured mouse mammary carcinoma FM3A cells. 631 74

We demonstrate that the activity of the major DNA topoisomerase I from calf thymus is severely inhibited after modification by purified poly(ADP-ribose) synthetase. Polymeric chains of poly(ADP-ribose) are covalently attached to DNA topoisomerase I. These observations with highly purified enzymes suggest that poly(ADP-ribosylation) may be a cellular mechanism for modulating DNA topoisomerase I activity in response to the state of DNA in the nucleus. Although extensive poly(ADP-ribosylation) of the Mr = 100,000 DNA topoisomerase I from calf thymus resulted in greater than 90% enzyme inhibition, exogenous poly(ADP-ribose) does not, by itself, inhibit topoisomerase activity. After modification, the apparent molecular weight of both the topoisomerase enzyme protein and of the topoisomerase enzyme activity was increased. In vitro, the extent of modification of DNA topoisomerase I could be controlled either by changing the ratio of topoisomerase to the synthetase or by varying the reaction time. More than 40 residues of ADP ribose per topoisomerase molecule could be added by the synthetase. Analysis of a poly(ADP-ribosylated) topoisomerase preparation that was about 50% inhibited revealed an average polymer chain length of 7.4, with 1-2 chains per enzyme molecule.
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PMID:Poly(ADP-ribosylation) of DNA topoisomerase I from calf thymus. 632 15

A water extract of Coptis chinensis was found to have the ability to stabilize the cleavable complex with mammalian DNA topoisomerase I. As the result of bioassay-guided fractionation, two protoberberine alkaloids, epiberberine and groenlandicine, were identified as active principles with topoisomerase I-mediated DNA cleavage activity in vitro. These two alkaloids did not induce topoisomerase II-mediated DNA cleavage. During further examination of the structurally related protoberberine alkaloids, berberrubine which is produced during the processing of Coptis rhizome as traditional medicine, was identified as a specific inducer of topoisomerase II-mediated DNA cleavage in vitro. These results indicated that protoberberine alkaloids are a chemical family which can induce cleavable complexes with topoisomerases I and II.
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PMID:Inhibitors of DNA topoisomerase I and II isolated from the Coptis rhizomes. 748 Feb 1


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