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At least 19 recordsLinked to original sources
PubMed1977

Ultraviolet light irradiation of PM2 superhelical DNA.

Superhelical PM2 DNA can be photochemically modified by u.v. irradiation. The variation of S20,w with dose shows the following characteristics. There is a linear increase from 28 to 31s produced by a low dose of u.v. irradiation (4,000 ergs/mm2). A plateau in S20,w occurs between 4,000 and 10,000 ergs/mm2. The S20,w then increases when irradiation is increased to 56,000 ergs/mm2. Thymine dimers are introduced proportional to dose throughtout the range of exposure to u.v. light. Sedimentation velocity-dye titrations reveal anomolous behavior, i.e. apparent increases in superhelix density (sigma). However, the dye-buoyant density procedure showed no change in sigma under the same conditions. The most satisfactory model for the data is preferential photochemical modification of premelted (possibly hairpin) sites as a greater rate than the introduction of photoproducts into duplex sites. The origin of the anomoly in the sedimentation velocity dye titrations is still unclear.

Bacteriophages↗
PubMedDate not supplied

[Effect of superhelical DNA on the transcription of cloned genes of the T4 phage].

The effect of DNA superhelicity on the transcription of T4 DNA fragments containing early genes uvs W, Y and late genes 25-29 was studied. RNA polymerase transcribes both early and late phage genes within the supercoiled recombinant plasmid. Late genes relative transcription increases essentially when T4-modified RNA polymerase is used. DNA relaxation causes a sharp decrease of modified RNA polymerase activity, especially on the late genes. The same effect is obtained in intact cells with recombinant plasmid, which superhelicity is lowered by temperature-sensitive mutation in DNA gyrase. The data obtained prove that DNA superhelicity is required for the T4 late transcription by phage-modified RNA polymerase. It is suggested that the dependence of late transcription on the phage DNA replication in infected cells is connected with phage DNA superspiralization throughout its replication.

Cloning, Molecular↗
PubMed1979

Superhelical DNA in Streptococcus sanguis: role in recombination in vivo.

Competent Streptococcus sanguis treated with non-lethal doses of coumermycin A1 immediately before or after uptake of radioactive transforming DNA were reduced in their capacity to yield transformants. This treatment did not alter bacterial ability to bind DNA in DNase I-resistant form, nor did it prevent the single-stranded donor DNA-recipient protein complexes formed upon uptake at the surface of the bacteria from translocating to chromosomal sites. Inhibition of transformation by heterospecific DNA was greater than that by homospecific DNA. The reduction in transformant yield was not accompanied by any loss of donor counts incorporated into the recipient chromosome, but rather by a loss of genetic activity of incorporated donor material indicating a failure of genetic integration and degradation of donor DNA as a consequence of coumermycin treatment. The inhibitory effect of coumermycin on transformation was associated with in vivo loss of chromosomal DNA superhelicity, The chromosomal DNA remained intact, however, indicative of inhibition of a gyrase-like enzyme responsible for the maintenance of negative supercoiling of the S. sanguis chromosome. Upon treatment with the drug, a coumermycin-resistant mutant strain showed neither loss of chromosomal superhelicity nor any inhibitory effect on genetic integration of donor DNA. The evidence supports the idea that chromosomal superhelicity promotes genetic recombination in vivo.

Aminocoumarins↗
PubMed1985

A pH-dependent structural transition in the homopurine-homopyrimidine tract in superhelical DNA.

We have inserted the 509-bp-long fragment of sea urchin P. miliaris histone gene spacer region into plasmid pUC19. The fragment contains the 60-bp-long homopurine-homopyrimidine tract that is known to be hypersensitive to the S1 endonuclease. Using two-dimensional gel electrophoresis we have observed a sharp structural transition in the insert with increasing DNA superhelicity. As in the cases of cruciform and Z form formation, the observed transition partly relaxes the superhelical stress. In contrast with the other two well documented transitions, the observed transition strongly depends on pH. At pH7 and above the transition occurs at negative superhelicities exceeding the physiological range (- sigma greater than 0.08). For pH6 the transition occurs at -sigma = 0.055, whereas for pH4.3 it takes place at -sigma = 0.001. A comprehensive analysis of the obtained data has made it possible to define the nature of the observed transition. We conclude that under superhelical stress or/and at low pH homopurinehomopyrimidine tracts adopt a novel spatial structure called the H form.

Animals↗
PubMed2000

[Thermo- and radioinduced enzymatic relaxation of superhelical DNA from mud loach Misgurnus fossilis L. spermatozoa].

Actions of environmental impacts on mud loach spermatozoa were studied using various model systems: a) temperature stress, b) X-ray irradiation in vivo only of the animal head (a condition to trigger stress reaction), c) X-ray irradiation in vivo only of the animal body (a condition to exclude a direct activation of principal stress-realizing organism systems), d) gamma-irradiation in vitro of the cell suspension. It has been demonstrated that the temperature stress or X-ray irradiation of the mud loach head induced three lines of effects: 1) significant decrease in DNA superhelical density, 2) activity redistribution (functional activation) of DNase II between chromatin subfractions (with the increase of its association to chromatin), and 3) intracellular acidification up to pH value to satisfy the DNase II initiation. The obtained facts allow to suggest that, first, DNase II participates in the presented temperature- and radio-induced supercoiled DNA relaxation in spermatozoa, and, second, DNase II is involved in physiological (season elimination of spermatozoa that remained within male gonads after fertilization) or environmentally-induced DNA degradation.

Animals↗
PubMed1990

The B-A transition in superhelical DNA.

Relaxation of a DNA superhelical stress due to the B to A transition induced by trifluoroethanol has been studied by assessing the change of DNA orientation in a flow gradient. Using DNAs of different superhelical densities, a decrease in the winding angle during the B----A shift of DNA was found to be 1.5 degrees per base pair in solution. Accepting the winding angle for B-DNA in solution to be 34.1 degrees, that for A-DNA must have a value of 32.6 degrees which agrees with the X-ray data for A-DNA in the condensed state. The date obtained within the B-A transition interval make it possible to conclude that there is an increase in winding at each B/A junction, which is about 5 degrees per one junction.

DNA Topoisomerases, Type I↗
PubMed1996

Prokaryotic DNA ligases unwind superhelical DNA.

We have studied the effect on DNA topology of binding of prokaryotic DNA ligases (T4 and E. coli) to superhelical or nicked circular DNA. Performing topoisomerase I-mediated relaxation in the presence of increasing amounts of T4 ligase led to a shift in the topoisomer distribution to increasingly more negative values. This result suggested that T4 ligase unwound the DNA and was further substantiated by ligation of nicked circular molecules by E. coli DNA ligase in the presence of increasing amounts of T4 ligase. Such an experiment was possible since the two DNA ligases require different cofactors for enzymatic activity. Performing a similar experiment with reverse partners, using E. coli DNA ligase as ligand, and T4 ligase as sealing agent, we observed that the E. coli enzyme also unwound the DNA. Thus, prokaryotic DNA ligases can be added to an ever-growing list of DNA-binding proteins that unwind the DNA upon binding.

Bacteriophage T4↗
PubMed1991

[An evaluation of the conformational changes in the superhelical DNA of eukaryotic cells by direct nucleoid fluorometry. II. The characteristics of the change in acridine orange fluorescence in studying the superhelical DNA of rat thymocytes].

Comparative studies of acridine orange (AO) and ethidium bromide interactions with supercoiled DNA (scDNA) of thymocytes were performed in which various conformational changes were induced. AO may be efficiently used for evaluation of conformational alterations of scDNA. Moreover, employing the maximum values of AO fluorescence allows to determine the maximum levels of scDNA relaxation.

Acridine Orange↗
PubMed1975

Interaction of f1 histone with superhelical DNA.

The superhelicity of double-stranded, closed circular SV40 DNA was altered by the addition of various amounts of ethidium bromide. The interaction of f1 histone with the series of molecules of various superhelicities was studied. The extent of interaction increases with increasing superhelicity regardless of whether it is of the positive or negative sense. The interaction of f1 histone with superhelical DNA is shown to be reversible.

Binding Sites↗
PubMed1983

[Nature of conformational restrictions in nuclear superhelical DNA from thymus lymphocytes].

The lymphocyte nucleoids of mouse thymus contain about 40% of rapidly labelled nuclear RNA, about 9% of total intracellular protein and all nuclear DNA. Relaxation of superhelical DNA after thymocyte nucleoids treatment with pronase or RNAase suggests that non-histone proteins and/or RNAs are involved in conformational restrictions in the superhelical domains of cell DNAs. Thymocyte nucleoids proteins are represented by two groups of nonhistone proteins with molecular weights of 50 000-60 000 and 75 000-85 000. An essential role in the appearance of conformational restrictions in thymocyte superhelical DNA belongs to disulfide bonds.

Animals↗
PubMed1981

Differences among subfractions of H1 histone in their interactions with linear and superhelical DNA. Circular dichroism.

Interactions between subfractions of ox thymus H1 histone and either linear T7 DNA or superhelical PM2 DNA were studied by measuring the circular dichroism of H1:DNA complexes. H1 subfractions differed from one to the next in their effectiveness at distorting the circular dichroic spectrum of DNA by as much as 3- to 4-fold for both forms of DNA. The order of effectiveness of the subfractions was the same at all ionic strengths between 0.05 M and 0.25 M, but the degree of spectral distortion caused by any of the subfractions was sensitive to the salt concentration. At 0 M NaCl and above 3 M NaCl, there was little or no distortion of the spectrum of DNA by any subfraction; the maximum effectiveness for all of the subfractions was at 0.15 M to 0.2 M NaCl whether the DNA was linear or superhelical. Between 0 M and 0.15 M NaCl, the H1 subfractions in free solution underwent a conformational change from a substantially unfolded state to one that is presumably the native state. This was revealed by circular dichroism. In part, this folding of the protein molecules must account for the effect of salt on the ability of H1 to distort the circular dichroism of DNA when the two macromolecules are brought together in complex formation. The distortion of the circular dichroism of DNA by H1:DNA complex formation is thought to be due to side-by-side aggregation of fibers in an asymmetrically ordered array. Apparently, the different H1 subfractions induce formation of H1:DNA complexes that differ in degree of orderliness or in a more complicated geometric parameter of the array, and this is true for superhelical as well as linear DNA.

Animals↗
PubMed1990

DNA structural polymorphism modulates the kinetics of superhelical DNA cleavage by BamHI restriction endonuclease.

A compartmental model developed by Hensley (Hensley, P., Nardone, G., Chirikjian, J.G., and Wastney, M. E., (1990) J. Biol. Chem. 265, 15300-15307) for analysis of the time courses of the cleavage of superhelical DNA substrates by the restriction endonuclease, BamHI, has been used to quantify the effects of changes in temperature, ionic strength, superhelical density, and the DNA substrate on the binding and strand cleavage processes. Studies reported here indicate that changes in topology may be introduced into the DNA substrate solely as a result of the plasmid preparation process and in the absence of covalent bond cleavage and ligation. These changes in topology have qualitatively different effects on the kinetics than those promoted by changes in the superhelical density. The former are removed by briefly warming the DNA prior to assay, suggesting that they are only kinetically stable, while the latter changes are not affected by heating. Increasing the [NaCl] from 0.01 M to 0.1 M increases the overall rate of plasmid cleavage by increasing both the rates of cleavage and enzyme DNA association. To describe the decrease in the overall cleavage rate observed in 0.15 M NaCl, an ionic strength-dependent rate-determining structural transition in the DNA substrate was incorporated into the model. The largest changes in the rate of the cleavage process resulted from changes in the DNA substrate. For the SV40 substrate compared to pBR322, the rate constants describing the two association processes and the first bond cleavage event were increased 6- to 7-fold. The rate of the second bond cleavage process was not affected. These changes may be due to differences in the flanking sequences.

DNA, Superhelical↗
PubMed1979

Purified Escherichia coli recA protein catalyzes homologous pairing of superhelical DNA and single-stranded fragments.

Purified Escherichia coli recA protein catalyzed ATP-dependent pairing of superhelical DNA and homologous single-stranded fragments. The product of the reaction: (i) was retained by nitrocellulose filters in 1.5 M NaCl/0.15 M Na citrate at pH 7, (ii) was dissociated at pH 12.3 but was not dissociated by heating at 55 degrees C for 4 min or by treatment with 0.2% sodium dodecyl sulfate and proteinase K, (iii) contained covalently closed circular double-stranded DNA (form I DNA), (iv) contained single-stranded fragments associated with replicative form (RF) DNA, and (v) contained a significant fraction of D-loops as judged by electron microscopy. Linear and nicked circular double-stranded DNA did not substitute well for superhelical DNA; intact circular single-stranded DNA did not substitute well for single-stranded fragments. Homologous combinations of single-stranded fragments and superhelical DNA from phages phiX174 and fd reacted, whereas heterologous combinations did not. The reaction required high concentrations of protein and MgCl2. The ATPase activity of purified recA protein was more than 98% dependent on the addition of single-stranded DNA. In 1 mM MgCl2, the ability of superhelical DNA to support the ATPase activity was two-thirds as good as that of single-stranded DNA.

Adenosine Triphosphatases↗
PubMedDate not supplied

[Enzymatic cleavage of superhelical DNA in a liquid crystal state].

Superhelical pBR322 DNA molecules form liquid-crystalline dispersions in water-salt solutions containing poly(ethyleneglycol). The formation of the liquid-crystalline dispersions from superhelical DNA molecules results in the appearance of two sites inside the DNA molecules that are split by Micrococcal nuclease. The first site of digestion does not differ from the standard site split by this enzyme in water-salt solutions, whereas the second one represents a new site specific only for the DNA molecules forming liquid-crystalline dispersions. Splitting of the DNA molecule through the first site is accompanied by formation of its linear form; splitting of a new site results in the formation of two linear DNA fragments with molecular masses equal to half of the initial DNA molecules. Enzyme digestion of superhelical DNA molecules forming liquid-crystalline dispersions induces a reformation of the "nonspecific" space organization of dispersions to the cholesteric one. A hypothetic model for packing of the superhelical DNA molecules inside liquid-crystalline dispersions and its transformation under enzyme action is suggested.

Circular Dichroism↗