Gene/Protein Disease Symptom Drug Enzyme Compound
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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)

Simian virus 40 (SV40) nucleoprotein complexes were studied with the electron microscope. Depending on the isolation procedure, SV40 chromatin has two different conformations: complexes isolated in the presence of 0.15 M NaCl appeared as very compact globular structures, while those isolated in the presence of 0.6 M NaCl had the typical 'beads-on-a-string' appearance of the primary nucleofilament. Concomitant with this structural change was a variation in the histone pattern and sedimentation behaviour of the complexes: with NaCl at 0.15 mol 1(-1) the isolated complexes contained both the nucleosomal histones and histone H1, and sedimented in sucrose gradients at 70S. Increasing the ionic strength to 0.6 M NaCl resulted in the removal of histone H1 from the complexes and in a decrease of the sedimentation coefficient to 40S. DNA relaxing enzyme is associated with the SV40 nucleoprotein complexes. The numbers of superhelical turns in DNA from compact and open types of complexes were found to be the same. Therefore the transition from the condensed to the open structure of viral chromatin does not require a change in the topological winding number of its DNA.
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PMID:The structure of SV 40 chromatin. 2 73

Extracts of Drosophila embryos can mediate the assembly of a chromatinlike structure from histones and DNA under physiological conditions. The histone-DNA complex formed in vitro contains micrococcal nuclease-sensitive sites spaced at 200-base pair intervals. More extensive digestion of the complex by micrococcal nuclease generates 11S particles which cosediment with nucleosome core particles isolated from native chromatin. These particles contain 140-base pair DNA fragments which upon further cleavage with micrococcal nuclease give rise to a pattern of discretely sized DNA fragments characteristic of nucleosome core particles. We have assayed the chromatin assembly process both qualitatively by measuring the induction of supertwists into a relaxed circular DNA (a process requiring a nicking-closing enzyme) and quantitatively by measuring the formation of micrococcal nuclease-resistant DNA fragments from radioactively labeled linear DNA. The amount of chromatin formed depends primarily on the amount of histones, whereas the rate of assembly depends on the amount of extract protein added. The factors in the extract that mediate chromatin assembly appear to interact first with the DNA because preincubation of the DNA with the extract markedly increases the extent of assembly.
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PMID:Extracts of Drosophila embryos mediate chromatin assembly in vitro. 11 49

The four core histones (H2A, H2B, H3, and H4) and DNA were assembled into nucleosome-like particles at physiological ionic strengths either by an extract of chromatin rich in nicking-closing activity or by the purified nicking-closing enzyme itself. When histone-DNA complexes were assembled in vitro from relaxed circular DNA, nearly physiological numbers of superhelical turns were induced in the DNA molecule. Electron microscopy of the complexes assembled by the chromatin extract revealed a beaded structure and a reduction of the contour length compared to free DNA. Micrococcal nuclease digestion of the histone-DNA complexes yielded 145-base-pair DNA fragments typical of nucleosome core particles and shorter subnucleosomal DNA fragments of discrete length.
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PMID:Nicking-closing enzyme assembles nucleosome-like structures in vitro. 22 80

We have formed complexes of relaxed closed circular Col E1 DNA with various combinations of histones, and examined the effects of treating the complexes with nicking-closing enzyme. Germond et al (1) have shown that when a mixture of the four core histones of the nucleosome (HIA, H2B, H3 and H4) is used in such an experiment, the subsequently isolated DNA is supercoiled. We find that the arginine-rich histone pair, H3 and H4, is sufficient to induce the supercoiling observed in this experiment. Both H3 and H4 are required, and in the absence of either, no other histones are effective. H3 and and H4 are as efficient, per unit weight, as a mixture of the four histones in inducing supercoils. We also show that there is a large difference between the DNA bending energy needed to form a nucleosome and that needed to form one turn of normal superhelical DNA. These two processes are energetically quite distinct and probably separable. We estimate the free energy of interaction between DNA-bound histone pairs, and find that one or two such interactions would generate enough energy to fold the DNA into a nucleosome.
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PMID:Supercoiling energy and nucleosome formation: the role of the arginine-rich histone kernel. 33 Dec 50

The action of DNA-relaxing enzyme on H1-DNA complexes was investigated. Complexes of superhelical and relaxed closed circular duplex DNA with H1 were treated with mammalian relaxing enzyme, deproteinized, and electrophoresed on agarose gels. At relatively low ratios of H1 to superhelical DNA, molecules of superhelical density intermediate between those of the starting material and relaxed DNA, the normal product, were generated. At relatively high H1 histone concentrations (H1:DNA greater than 0.4 w/w), the superhelical DNA was not relaxed. Further, no superhelical turns were introduced into relaxed closed duplex DNA at any concentration of H1 tested. Thus, the binding of H1 histone to DNA prevents the action of the relaxing enzyme. Moreover, H1 histone does not appear to unwind the DNA duplex upon binding. The implications of these observations and the previously demonstrated specificity of H1 histone for superhelical DNA are discussed in relation to the structure of chromatin.
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PMID:The effect of H1 histone on the action of DNA-relaxing enzyme. 86 71

The interaction of closed circular duplex DNA with the lysine-rich H5 histone fraction of avian erythrocytes has been studied. H5, like H1 histone, interacts preferentially with superhelical DNA. The extent of interaction increases with increasing negative or positive superhelicity. Salt-extracted lysine-rich histones show the same specificity for interaction with superhelices as do acid-extracted preparations. Chicken erythrocyte nuclei contain DNA-relaxing enzyme. This enzyme is extracted from the nuclei at lower salt concentrations than those required to extract H1 and H5 histones and is, therefore, probably a function of a protein distinct from H1 and H5 histones.
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PMID:H5 Histone and DNA-relaxing enzyme of chicken erythrocytes. Interaction with superhelical DNA. 100 94

We have identified two classes of in vivo topoisomerase II cleavage sites in the Drosophila histone gene repeat. One class co-localizes with DNase I-hypersensitive regions and another novel class maps to a subset of consecutive nucleosome linker sites in the scaffold-associated region (SAR) of the histone gene loop. Prominent topoisomerase II cleavage is also observed in one of the linker regions of the two nucleosomes spanning satellite III, a centromeric SAR-like DNA sequence with a repeat length of 359 bp. At the sequence level, in vivo topoisomerase II cleavage is highly site specific. Comparison of 10 nucleosome linker sites defines an in vivo cleavage sequence whose major characteristic is a prominent GC-rich core. These GC-rich cleavage sites are flanked by extensive arrays of oligo(dA).oligo(dT) tracts characteristic of SAR sequences. Treatment of cells with distamycin selectively enhances cleavage at nucleosome linker sites of the SAR and satellite regions, suggesting that AT-rich sequences flanking cleavage sites may be involved in determining topoisomerase II activity in the cell. These observations provide evidence for the association of topoisomerase II with SARS in vivo.
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PMID:In vivo topoisomerase II cleavage of the Drosophila histone and satellite III repeats: DNA sequence and structural characteristics. 131 Dec 55

In a variety of adult and childhood leukaemia cell samples collected at different states of the disease, we analysed in a series of sequentially performed slot-blot or Northern-blot hybridisation experiments the expression of genes possibly involved in multiple drug resistance (MDR) (mdr1/P-glycoprotein, DNA topoisomerase II, glutathione-S-transferase pi), and the expression of the DNA topoisomerase I and histone 3.1 genes. Occasionally, P-glycoprotein gene expression was additionally examined by indirect immunocytofluorescence using the monoclonal antibody C219. No significant difference in mdr1/P-glycoprotein mRNA levels between primary and relapsed state acute lymphocytic leukaemias (ALL) was seen on average. Second or third relapses, however, showed a distinct tendency to an elevated expression of this multidrug transporter gene (up to 10-fold) in part well beyond the value seen in the moderately cross-resistant T-lymphoblastoid CCRF-CEM subline CCRF VCR 100. Increased mdr1/P-glycoprotein mRNA levels were also found in relapsed state acute myelogenous leukaemias (AML), and in chronic lymphocytic leukaemias (CLL) treated with chlorambucil and/or prednisone for several years. Topoisomerase I and topoisomerase II mRNA levels were found to be very variable. Whereas in all but one case of CLL topoisomerase II mRNA was not detected by slot-blot hybridizations, strong topoisomerase I and topoisomerase II gene expression levels, frequently exceeding the levels monitored in the CCRF-CEM cell line, were seen in many cell samples of acute leukaemia. If topoisomerase II mRNA was undetectable, expression of topoisomerase I was clearly visible throughout. These observations might be valuable considering the possible treatment with specific topoisomerase I or topoisomerase II inhibitors. Significant positive correlations were found (i) for topoisomerase I and histone 3.1 gene expression levels in general (P less than 0.001), and (ii) in the CLL samples additionally for the expression levels of the mdr1 gene, and the histone 3.1, topoisomerase I, and glutathione-S-transferase pi genes, respectively.
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PMID:Mdr1/P-glycoprotein, topoisomerase, and glutathione-S-transferase pi gene expression in primary and relapsed state adult and childhood leukaemias. 135 60

In the studies reported here we have used topoisomerase II as a model system for analyzing the factors that determine the sites of action for DNA-binding proteins in vivo. To localize topoisomerase II sites in vivo we used an inhibitor of the purified enzyme, the antitumor drug VM-26. This drug stabilizes an intermediate in the catalytic cycle, the cleavable complex, and substantially stimulates DNA cleavage by topoisomerase II. We show that lysis of VM-26 treated tissue culture cells with sodium dodecyl sulfate induces highly specific double-strand breaks in genomic DNA, and we present evidence indicating that these double-strand breaks are generated by topoisomerase II. Using indirect end labeling to map the cleavage products, we have examined the in vivo sites of action of topoisomerase II in the 87A7 heat shock locus, the histone repeat, and a tRNA gene cluster at 90BC. Our analysis reveals that chromatin structure, not sequence specificity, is the primary determinant in topoisomerase II site selection in vivo. We suggest that chromatin organization may provide a general mechanism for generating specificity in a wide range of DNA-protein interactions in vivo.
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PMID:Chromatin structure, not DNA sequence specificity, is the primary determinant of topoisomerase II sites of action in vivo. 165 19

Some models of in vitro chromatin assembly suggest a biphasic molecular mechanism. The first phase, nucleosome formation, is comprised of the formation of histone-DNA complexes which mature into a canonical nucleosome structure. The second phase represents the process by which these nucleosomes become properly spaced with a regular periodicity on the DNA. In this report, we examine the role of DNA topoisomerases in the latter phase of chromatin assembly. To study this process, we use a Xenopus laevis cell-free extract, which assembles quantitative amounts of chromatin on circular DNA templates, and the type II topoisomerase-specific antitumor drugs VM-26 and endrofloxicin. Our results suggest that nucleosome formation is unaffected by the presence of VM-26 or endrofloxicin. However, periodic spacing of nucleosomes is inhibited significantly by these drugs. In the absence of proper chromatin assembly, circular DNA molecules are processed into nucleoprotein complexes which are transcribed poorly. Taken together, these results indicate that the antitumor drugs VM-26 and endrofloxicin influence gene expression indirectly by blocking the periodic spacing of nucleosomes.
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PMID:Assembly of transcriptionally active chromatin in vitro: a possible role for topoisomerase II. 196 75


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