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Nicholas R. Cozzarelli - One of the best experts on this subject based on the ideXlab platform.
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alteration of escherichia coli topoisomerase iv conformation upon enzyme binding to positively Supercoiled DNA
Journal of Biological Chemistry, 2006Co-Authors: Nancy J Crisona, Nicholas R. CozzarelliAbstract:Abstract Escherichia coli topoisomerase IV (topo IV) is an essential enzyme that unlinks the daughter chromosomes for proper segregation at cell division. In vitro, topo IV readily distinguishes between the two possible chiralities of crossing segments in a DNA substrate. The enzyme relaxes positive supercoils and left-handed braids 20 times faster, and with greater processivity, than negative supercoils and right-handed braids. Here, we used chemical cross-linking of topo IV to demonstrate that enzyme bound to positively Supercoiled DNA is in a different conformation from that bound to other forms of DNA. Using three different reagents, we observed novel cross-linked species of topo IV when positively Supercoiled DNA was in the reaction. We show that the ParE subunits are in close enough proximity to be cross-linked only when the enzyme is bound to positively Supercoiled DNA. We suggest that the altered conformation reflects efficient binding by topo IV of the two DNA segments that participate in the strand passage reaction.
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the effect of ionic conditions on the conformations of Supercoiled DNA ii equilibrium catenation
Journal of Molecular Biology, 1997Co-Authors: Valentin V. Rybenkov, Alexander Vologodskii, Nicholas R. CozzarelliAbstract:We studied the equilibrium formation of DNA catenanes to assess the conformational properties of Supercoiled DNA as a function of ionic conditions and supercoiling density. Catenanes were formed by cyclizing linear DNA with long cohesive ends in the presence of Supercoiled molecules. The efficiency of the catenation depends on the distance between opposing segments of DNA in the interwound superhelix. The fraction of cyclizing molecules that becomes topologically linked with the Supercoiled DNA is the product of the concentration of the Supercoiled DNA and a proportionality constant, B, that depends on conformations of Supercoiled DNA. In parallel with these experimental studies, we calculated the values of B using Monte Carlo simulations of the equilibrium distribution of DNA conformations. There were no adjustable parameters in the calculations because all three parameters of the DNA model, bending and torsional elasticity of DNA and DNA effective diameter, specifying intersegment interactions, were known from independent studies. We found very good agreement between measured and simulated values of B for all the ionic conditions and DNA superhelix densities studied; the discrepancy was less than a factor of 2 over the 200-fold variation in B. The value of B decreases nearly exponentially with increasing superhelicity, this dependence being especially strong at low salt concentration. The dependence of B on the concentration of NaCl, MgCl2, and spermidine can be described with good accuracy in terms of changes of the DNA effective diameter. We found no indication of superhelix collapse under any ionic conditions studied. We discuss, in light of these results, the biological importance of the effect of DNA supercoiling on the unlinking of the products of DNA replication. # 1997 Academic Press Limited
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the effect of ionic conditions on the conformations of Supercoiled DNA i sedimentation analysis
Journal of Molecular Biology, 1997Co-Authors: Valentin V. Rybenkov, Alexander Vologodskii, Nicholas R. CozzarelliAbstract:We studied the conformations of Supercoiled DNA as a function of superhelicity and ionic conditions by determining its sedimentation coefficient both experimentally and by calculation. To cancel out unknown parameters from both calculations and experiments, we determined the ratio of the sedimentation coefficient, s, to that of open circular DNA, soc. Calculations of the sedimentation coefficient were based on direct solution of the Burgers-Oseen problem for an equilibrium set of DNA conformations generated for each condition by the Metropolis Monte Carlo procedure. There were no adjustable parameters in the Monte Carlo simulations because all three parameters of the DNA model used, bending and torsional elasticity of DNA and DNA effective diameter specifying electrostatic interactions, were known from independent data. The good agreement between measured and calculated values of s/soc allowed us to interpret the sedimentation results in terms of DNA conformations, with particular emphasis on the marked effect of ionic conditions. As NaCl concentration decreases, s/soc increases because the superhelix becomes less regular and more compact. In the presence of just 10 mM MgCl2, Supercoiled DNA adopts essentially the same set of conformations as in moderate to high concentrations of NaCl. Our simulations showed that s is a strong function of the superhelix branching frequency. At near physiological ionic conditions, there are about four branches in the 7 kb DNA molecule used in this work. We found no indication of superhelix collapse in any ionic conditions even remotely approaching physiological ones. For all ionic conditions studied, we conclude that the electrostatic interaction of DNA segments specified by the DNA effective diameter is the primary determinant of Supercoiled DNA conformations. # 1997 Academic Press Limited
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The effect ionic conditions on the conformations of Supercoiled DNA. I. Sedimentation analysis
Journal of Molecular Biology, 1997Co-Authors: Valentin V. Rybenkov, Alexander V. Vologodskii, Nicholas R. CozzarelliAbstract:We studied the conformations of Supercoiled DNA as a function of superhelicity and ionic conditions by determining its sedimentation coefficient both experimentally and by calculation. To cancel out unknown parameters from both calculations and experiments, we determined the ratio of the sedimentation coefficient, s, to that of open circular DNA, s(oc). Calculations of the sedimentation coefficient were based on direct solution of the Burgers-Oseen problem for an equilibrium set of DNA conformations generated for each condition by the Metropolis Monte Carlo procedure. There were no adjustable parameters in the Monte Carlo simulations because all three parameters of the DNA model used, bending and torsional elasticity of DNA and DNA effective diameter specifying electrostatic interactions, were known from independent data. The good agreement between measured and calculated values of s/s(oc) allowed us to interpret the sedimentation results in terms of DNA conformations, with particular emphasis on the marked effect of ionic conditions. As NaCl concentration decreases, s/s(oc) increases because the superhelix becomes less regular and more compact. In the presence of just 10 mM MgCl2, Supercoiled DNA adopts essentially the same set of conformations as in moderate to high concentrations of NaCl. Our simulations showed that s is a strong function of the superhelix branching frequency. At near physiological ionic conditions, there are about four branches in the 7 kb DNA molecule used in this work. We found no indication of superhelix collapse in any ionic conditions even remotely approaching physiological ones. For all ionic conditions studied, we conclude that the electrostatic interaction of DNA segments specified by the DNA effective diameter is the primary determinant of Supercoiled DNA conformations.
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conformational and thermodynamic properties of Supercoiled DNA
Annual Review of Biophysics and Biomolecular Structure, 1994Co-Authors: Alexander Vologodskii, Nicholas R. CozzarelliAbstract:: Work in the 1990s has substantially increased our understanding of supercoiling conformations and energetics. We now know many of the basic properties of Supercoiled DNA as a result of the synergy between experimental and theoretical analyses. We conclude by summarizing the results. 1. All available data indicate a plectonemic structure for Supercoiled DNA. First, three types of EM (conventional, cryo, and scanning force) show the plectonemic form. Second, the topology of the catenanes and knots generated from Supercoiled DNA by the Int recombinase demands that the substrate supercoils are plectonemic, as does the topology of knotting by type-2 topoisomerases. Third, all the theoretical and computer analyses indicate that the superhelix has the interwound form. 2. The superhelix conformations are often branched, as observed using EM and Monte Carlo simulation. Moreover, branching is required to explain the distribution of knots and catenanes produced by Int or topoisomerases as well as the dependence of s and Rg on sigma. Branching frequency is very sensitive to sigma, DNA length, ionic conditions, DNA bends, and temperature. Despite the qualitative agreement, the quantitative differences between experimental and computational data point out the need for further studies of branching. 3. The results of Monte Carlo simulations, theoretical analyses, and cryo-EM show that the conformational and thermodynamic properties of Supercoiled DNA depend strongly on ionic conditions. The reason for such a dependence is clear. Counterions shield DNA negative charges and decrease the repulsion of DNA segments in the tight interwound structure. The effective double-helix diameter increases from 3 to 15 nm as the salt concentration is reduced from 1.00 to 0.01 M. Experimental investigations of the dependence on ionic conditions of Supercoiled DNA properties are just beginning. The following conclusions refer to conditions of moderate to high monovalent or divalent ion concentrations (> or = 0.1 M [Na+] or > or = 0.01 M [Mg2+], respectively). 1. Wr takes up about 3/4 of the delta Lk, and Wr/delta Tw is independent of sigma for DNA > or = 2.5 kb in length. A constant ratio is implied by the CD data, and Wr/delta Tw has been determined with conventional EM, cryo-EM, the Int topological method, and Monte Carlo simulation. 2. The average number of supercoils is (0.8 to 0.9) x delta Lk and is independent of DNA length. These results were obtained with EM, the Int topological method, and computer simulation.(ABSTRACT TRUNCATED AT 400 WORDS)
Alexander Vologodskii - One of the best experts on this subject based on the ideXlab platform.
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Unlinking of Supercoiled DNA Catenanes by Type IIA Topoisomerases
Biophysical Journal, 2011Co-Authors: Alexander VologodskiiAbstract:It was found recently that DNA catenanes, formed during replication of circular plasmids, become positively (+) Supercoiled, and the unlinking of such catenanes by type IIA topoisomerases proceeds much more efficiently than the unlinking of negatively (−) Supercoiled catenanes. In an attempt to explain this striking finding we studied, by computer simulation, conformational properties of Supercoiled DNA catenanes. Although the simulation showed that conformational properties of (+) and (−) Supercoiled replication catenanes are very different, these properties per se do not give any advantage to (+) Supercoiled over (−) Supercoiled DNA catenanes for unlinking. An advantage became evident, however, when we took into account the established features of the enzymatic reaction catalyzed by the topoisomerases. The enzymes create a sharp DNA bend in the first bound DNA segment and allow for the transport of the second segment only from inside the bend to its outside. We showed that in (−) Supercoiled DNA catenanes this protein-bound bent segment becomes nearly inaccessible for segments of the other linked DNA molecule, inhibiting the unlinking.
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dynamics of site juxtaposition in Supercoiled DNA
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Jing Huang, Tamar Schlick, Alexander VologodskiiAbstract:Juxtaposition kinetics between specific sites in Supercoiled DNA is investigated at close to physiological ionic conditions by Brownian dynamics simulations. At such conditions, Supercoiled DNA is interwound, and the probability of spatial site juxtaposition is much higher than in relaxed DNA. We find, however, that supercoiling does not correspondingly increase the rate of juxtaposition at these physiological conditions. An explanation to this unexpected finding emerges on analysis of the juxtaposition dynamics. We note that although a particular site i1 in Supercoiled DNA is often in close proximity (juxtaposed) to another site i2, the change of i2 occurs very slowly and depends largely on internal slithering of opposite segments of the DNA superhelix. Such slithering results in long correlations between successive values of i2; these correlations increase the average time of juxtaposition between two DNA sites. Random collisions between sites located on different superhelix branches—although increasing in importance with DNA size—contribute less substantially to site juxtaposition at high salt than slithering for DNA up to 6 kb in length.
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Effect of Magnesium on Cruciform Extrusion in Supercoiled DNA
Journal of Molecular Biology, 1999Co-Authors: Maria Vologodskaia, Alexander VologodskiiAbstract:Abstract Recently, it was reported that Mg2+greatly facilitates cruciform extrusion in the short palindromes of Supercoiled DNA, thereby allowing the formation of cruciform structures in vivo. Because of the potential biological importance of this phenomenon, we undertook a broader study of the effect of Mg2+on a cruciform extrusion in Supercoiled DNA. The method of two-dimensional gel electrophoresis was used to detect the cruciform extrusion both in the absence and in the presence of these ions. Our results show that Mg2+shifts the cruciform extrusion in the d(CCC(AT)16GGG) palindrome to a higher, rather than to a lower level of supercoiling. In order to study possible sequence-specific properties of the short palindromes for which the unusual cruciform extrusion in the presence Mg2+was reported, we constructed a plasmid with a longer palindromic region. This region bears the same sequences in the hairpin loops and four-arm junction as the short palindrome, except that the short stems of the hairpins are extended. The extension allowed us to overcome the limitation of our experimental approach which cannot be used for very short palindromes. Our results show that Mg2+also shifts the cruciform extrusion in this palindrome to a higher level of supercoiling. These data suggest that cruciform extrusion in the short palindromes at low supercoiling is highly improbable. We performed a thermodynamic analysis of the effect of Mg2+on cruciform extrusion. The treatment accounted for the effect of Mg2+on both free energy of supercoiling and the free energy of cruciform structure per se. Our analysis showed that although the level of supercoiling required for the cruciform extrusion is not reduced by Mg2+, the ions reduce the free energy of the cruciform structure.
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internal motion of Supercoiled DNA brownian dynamics simulations of site juxtaposition
Journal of Molecular Biology, 1998Co-Authors: Hongmei Jian, Tamar Schlick, Alexander VologodskiiAbstract:Thermal motions in Supercoiled DNA are studied by Brownian dynamics (BD) simulations with a focus on the site juxtaposition process. It had been shown in the last decade that the BD approach is capable of describing actual times of large-scale DNA motion. The bead model of DNA used here accounts for bending and torsional elasticity as well as the electrostatic repulsion among DNA segments. The hydrodynamic interaction among the beads of the model chain and the aqueous solution is incorporated through the Rotne-Prager tensor. All simulations were performed for the sodium ion concentration of 0.01 M. We first showed, to test our BD procedure, that the same distributions of equilibrium conformational properties are obtained as by Monte Carlo simulations for the corresponding DNA model. The BD simulations also predict with accuracy published experimental values of the diffusion coefficients of Supercoiled DNA. To describe the rate of conformational changes, we also calculated the autocorrelation functions for the writhe and radius of gyration for the Supercoiled molecules. The rate of site juxtaposition was then studied for DNA molecules up to 3000 bp in length. We find that site juxtaposition is a very slow process: although accelerated by a factor of more than 100 by DNA supercoiling, the times of juxtaposition are in the range of ms even for highly Supercoiled DNA, about two orders of magnitude higher than the relaxation times of writhe and the radius of gyration for the same molecules. By inspecting successive simulated conformations of Supercoiled DNA, we conclude that slithering of opposing segments of the interwound superhelix is not an efficient mechanism to accomplish site juxtaposition, at least for conditions of low salt concentration. Instead, transient distortions of the interwound superhelix, followed by continuous reshaping of the molecule, contribute more significantly to site juxtaposition kinetics. # 1998 Academic Press
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the effect of ionic conditions on the conformations of Supercoiled DNA ii equilibrium catenation
Journal of Molecular Biology, 1997Co-Authors: Valentin V. Rybenkov, Alexander Vologodskii, Nicholas R. CozzarelliAbstract:We studied the equilibrium formation of DNA catenanes to assess the conformational properties of Supercoiled DNA as a function of ionic conditions and supercoiling density. Catenanes were formed by cyclizing linear DNA with long cohesive ends in the presence of Supercoiled molecules. The efficiency of the catenation depends on the distance between opposing segments of DNA in the interwound superhelix. The fraction of cyclizing molecules that becomes topologically linked with the Supercoiled DNA is the product of the concentration of the Supercoiled DNA and a proportionality constant, B, that depends on conformations of Supercoiled DNA. In parallel with these experimental studies, we calculated the values of B using Monte Carlo simulations of the equilibrium distribution of DNA conformations. There were no adjustable parameters in the calculations because all three parameters of the DNA model, bending and torsional elasticity of DNA and DNA effective diameter, specifying intersegment interactions, were known from independent studies. We found very good agreement between measured and simulated values of B for all the ionic conditions and DNA superhelix densities studied; the discrepancy was less than a factor of 2 over the 200-fold variation in B. The value of B decreases nearly exponentially with increasing superhelicity, this dependence being especially strong at low salt concentration. The dependence of B on the concentration of NaCl, MgCl2, and spermidine can be described with good accuracy in terms of changes of the DNA effective diameter. We found no indication of superhelix collapse under any ionic conditions studied. We discuss, in light of these results, the biological importance of the effect of DNA supercoiling on the unlinking of the products of DNA replication. # 1997 Academic Press Limited
Andrzej Stasiak - One of the best experts on this subject based on the ideXlab platform.
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how topoisomerase iv can efficiently unknot and decatenate negatively Supercoiled DNA molecules without causing their torsional relaxation
Nucleic Acids Research, 2016Co-Authors: Eric J Rawdon, Julien Dorier, Dusan Racko, Kenneth C Millett, Andrzej StasiakAbstract:Freshly replicated DNA molecules initially form multiply interlinked right-handed catenanes. In bacteria, these catenated molecules become Supercoiled by DNA gyrase before they undergo a complete decatenation by topoisomerase IV (Topo IV). Topo IV is also involved in the unknotting of Supercoiled DNA molecules. Using Metropolis Monte Carlo simulations, we investigate the shapes of Supercoiled DNA molecules that are either knotted or catenated. We are especially interested in understanding how Topo IV can unknot right-handed knots and decatenate right-handed catenanes without acting on right-handed plectonemes in negatively Supercoiled DNA molecules. To this end, we investigate how the topological consequences of intersegmental passages depend on the geometry of the DNA-DNA juxtapositions at which these passages occur. We observe that there are interesting differences between the geometries of DNA-DNA juxtapositions in the interwound portions and in the knotted or catenated portions of the studied molecules. In particular, in negatively Supercoiled, multiply interlinked, right-handed catenanes, we detect specific regions where DNA segments belonging to two freshly replicated sister DNA molecules form left-handed crossings. We propose that, due to its geometrical preference to act on left-handed crossings, Topo IV can specifically unknot Supercoiled DNA, as well as decatenate postreplicative catenanes, without causing their torsional relaxation.
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The apical localization of transcribing RNA polymerases on Supercoiled DNA prevents their rotation around the template.
The EMBO Journal, 1992Co-Authors: B Ten Heggeler-bordier, Marc Adrian, Walter Wahli, Andrzej Stasiak, Jacques DubochetAbstract:Abstract The interaction of Escherichia coli RNA polymerase with Supercoiled DNA was visualized by cryo-electron microscopy of vitrified samples and by classical electron microscopy methods. We observed that when E. coli RNA polymerase binds to a promoter on Supercoiled DNA, this promoter becomes located at an apical loop of the interwound DNA molecule. During transcription RNA polymerase shifts the apical loop along the DNA, always remaining at the top of the moving loop. This relationship between RNA polymerase and the Supercoiled template precludes circling of the RNA polymerase around the DNA and prevents the growing RNA transcript from becoming entangled with the template DNA.
Tamar Schlick - One of the best experts on this subject based on the ideXlab platform.
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dynamics of site juxtaposition in Supercoiled DNA
Proceedings of the National Academy of Sciences of the United States of America, 2001Co-Authors: Jing Huang, Tamar Schlick, Alexander VologodskiiAbstract:Juxtaposition kinetics between specific sites in Supercoiled DNA is investigated at close to physiological ionic conditions by Brownian dynamics simulations. At such conditions, Supercoiled DNA is interwound, and the probability of spatial site juxtaposition is much higher than in relaxed DNA. We find, however, that supercoiling does not correspondingly increase the rate of juxtaposition at these physiological conditions. An explanation to this unexpected finding emerges on analysis of the juxtaposition dynamics. We note that although a particular site i1 in Supercoiled DNA is often in close proximity (juxtaposed) to another site i2, the change of i2 occurs very slowly and depends largely on internal slithering of opposite segments of the DNA superhelix. Such slithering results in long correlations between successive values of i2; these correlations increase the average time of juxtaposition between two DNA sites. Random collisions between sites located on different superhelix branches—although increasing in importance with DNA size—contribute less substantially to site juxtaposition at high salt than slithering for DNA up to 6 kb in length.
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internal motion of Supercoiled DNA brownian dynamics simulations of site juxtaposition
Journal of Molecular Biology, 1998Co-Authors: Hongmei Jian, Tamar Schlick, Alexander VologodskiiAbstract:Thermal motions in Supercoiled DNA are studied by Brownian dynamics (BD) simulations with a focus on the site juxtaposition process. It had been shown in the last decade that the BD approach is capable of describing actual times of large-scale DNA motion. The bead model of DNA used here accounts for bending and torsional elasticity as well as the electrostatic repulsion among DNA segments. The hydrodynamic interaction among the beads of the model chain and the aqueous solution is incorporated through the Rotne-Prager tensor. All simulations were performed for the sodium ion concentration of 0.01 M. We first showed, to test our BD procedure, that the same distributions of equilibrium conformational properties are obtained as by Monte Carlo simulations for the corresponding DNA model. The BD simulations also predict with accuracy published experimental values of the diffusion coefficients of Supercoiled DNA. To describe the rate of conformational changes, we also calculated the autocorrelation functions for the writhe and radius of gyration for the Supercoiled molecules. The rate of site juxtaposition was then studied for DNA molecules up to 3000 bp in length. We find that site juxtaposition is a very slow process: although accelerated by a factor of more than 100 by DNA supercoiling, the times of juxtaposition are in the range of ms even for highly Supercoiled DNA, about two orders of magnitude higher than the relaxation times of writhe and the radius of gyration for the same molecules. By inspecting successive simulated conformations of Supercoiled DNA, we conclude that slithering of opposing segments of the interwound superhelix is not an efficient mechanism to accomplish site juxtaposition, at least for conditions of low salt concentration. Instead, transient distortions of the interwound superhelix, followed by continuous reshaping of the molecule, contribute more significantly to site juxtaposition kinetics. # 1998 Academic Press
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Solvent effects on Supercoiled DNA dynamics explored by Langevin dynamics simulations.
Physical Review E, 1995Co-Authors: Gomathi Ramachandran, Tamar SchlickAbstract:The dynamical effects of solvent on Supercoiled DNA are explored through a simple, macroscopic energy model for DNA in the Langevin dynamics framework. Closed circular DNA is modeled by B splines, and both eleastic and electrostatic (screened Coulomb) potentials are included in the energy function. The Langevin formalism describes approximately the influence of the solvent on the motion of the solute. The collision frequency \ensuremath{\gamma} determines the magnitude of the friction and the variance of the random forces due to molecular collisions. Thus, as a first approximation, the Langevin equation of motion can be parametrized to capture the approximate dynamics of DNA in a viscous medium. Solvent damping is well known to alter the dynamical behavior of DNA and affect various hydrodynamic properties. This work examines these effects systematically by varying the collision frequency (viscosity) with the goal of better understanding the dynamical behavior of Supercoiled DNA. By varying \ensuremath{\gamma} over ten orders of magnitude, we identify three distinct physical regimes of DNA behavior: (i) low \ensuremath{\gamma}, dominated by globally harmonic motion; (ii) intermediate \ensuremath{\gamma}, characterized by maximal sampling and high mobility of the DNA; and (iii) high \ensuremath{\gamma}, dominated by random forces, where all of the global modes are effectively frozen by extreme overdamping. These regimes are explored extensively by Langevin dynamics simulations, offering insight into hydrodynamic effects on Supercoiled DNA. At low \ensuremath{\gamma}, the DNA exhibits small, harmonic fluctuations.
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The influence of salt on the structure and energetics of Supercoiled DNA.
Biophysical Journal, 1994Co-Authors: Tamar Schlick, Bin Li, Wilma K. OlsonAbstract:We present a detailed computational study of the influence of salt on the configurations, energies, and dynamics of Supercoiled DNA. A potential function that includes both elastic and electrostatic energy components is employed. Specifically, the electrostatic term, with salt-dependent coefficients, is modeled after Stigter's pioneering work on the effective diameter of DNA as a function of salt concentration. Because an effective charge per unit length is used, the electrostatic formulation does not require explicit modeling of phosphates and can be used to study long DNAs at any desired resolution of charge. With explicit consideration of the electrostatic energy, an elastic bending constant corresponding to the nonelectrostatic part of the bending contribution to the persistence length is used. We show, for a series of salt concentrations ranging from 0.005 to 1.0 M sodium, how configurations and energies of Supercoiled DNA (1000 and 3000 base pairs) change dramatically with the simulated salt environment. At high salt, the DNA adopts highly compact and bent interwound states, with the bending energy dominating over the other components, and the electrostatic energy playing a minor role in comparison to the bending and twisting terms. At low salt, the DNA supercoils are much more open and loosely interwound, and the electrostatic components are dominant. Over the range of three decades of salt examined, the electrostatic energy changes by a factor of 10. The buckling transition between the circle and figure-8 is highly sensitive to salt concentration: this transition is delayed as salt concentration decreases, with a particularly sharp increase below 0.1 M. For example, for a bending-to-twisting force constant ratio of A/C = 1.5, the linking number difference (delta LK) corresponding to equal energies for the circle and figure-8 increases from 2.1 to 3.25 as salt decreases from 1.0 to 0.005 M. We also present in detail a family of three-lobed Supercoiled DNA configurations that are predicted by elasticity theory to be stable at low delta Lk. To our knowledge, such three-dimensional structures have not been previously presented in connection with DNA supercoiling. These branched forms have a higher bending energy than the corresponding interwound configurations at the same delta Lk but, especially at low salt, this bending energy difference is relatively small in comparison with the total energy, which is dominated by the electrostatic contributions. Significantly, the electrostatic energies of the three-lobed and (straight) interwound forms are comparable at each salt environment.(ABSTRACT TRUNCATED AT 400 WORDS)
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On higher buckling transitions in Supercoiled DNA.
Biopolymers, 1994Co-Authors: Tamar Schlick, Wilma K. Olson, Timothy P. Westcott, Jerry P. GreenbergAbstract:: A combination of detailed energy minimization and molecular dynamics studies of closed circular DNA offers here new information that may be relevant to the dynamics of short DNA chains and/or low superhelical densities. We find a complex dependence of Supercoiled DNA energies and geometries on the linking number difference delta Lk as physiological superhelical densities (magnitude of sigma approximately 0.06) are approached. The energy minimization results confirm and extend predictions of classical elasticity theory for the equilibria of elastic rods. The molecular dynamics results suggest how these findings may affect the dynamics of Supercoiled DNA. The minimization reveals sudden higher order configurational transitions in addition to the well-known catastrophic buckling from the circle to the figure-8. The competition among the bending, twisting, and self-contact forces leads to different families of Supercoiled forms. Some of those families begin with configurations of near-zero twist. This offers the intriguing possibility that nicked DNA may relax to low-twist forms other than the circle, as generally assumed. Furthermore, for certain values of delta Lk, more than one interwound DNA minimum exists. The writhing number as a function of delta Lk is discontinuous in some ranges; it exhibits pronounced jumps as delta Lk is increased from zero, and it appears to level off to a characteristic slope only at higher values of delta Lk. These findings suggest that Supercoiled DNA may undergo systematic rapid interconversions between different minima that are both close in energy and geometry. Our molecular dynamics simulations reveal such transitional behavior. We observe the macroscopic bending and twisting fluctuations of interwound forms about the global helix axis as well as the end-over-end tumbling of the DNA as a rigid body. The overall mobility can be related to magnitude of sigma and to the bending, twisting, and van der Waals energy fluctuations. The general character of molecular motions is thus determined by the types of energy minima found at a given delta Lk. Different time scales may be attributed to each type of motion: The overall chain folding occurs on a time scale almost an order of magnitude faster than the end-over-end tumbling. The local bending and twisting of individual chain residues occur at an even faster rate, which in turn correspond to several cycles of local variations for each large-scale bending and straightening motion of the DNA.
Jacques Dubochet - One of the best experts on this subject based on the ideXlab platform.
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The apical localization of transcribing RNA polymerases on Supercoiled DNA prevents their rotation around the template.
The EMBO Journal, 1992Co-Authors: B Ten Heggeler-bordier, Marc Adrian, Walter Wahli, Andrzej Stasiak, Jacques DubochetAbstract:Abstract The interaction of Escherichia coli RNA polymerase with Supercoiled DNA was visualized by cryo-electron microscopy of vitrified samples and by classical electron microscopy methods. We observed that when E. coli RNA polymerase binds to a promoter on Supercoiled DNA, this promoter becomes located at an apical loop of the interwound DNA molecule. During transcription RNA polymerase shifts the apical loop along the DNA, always remaining at the top of the moving loop. This relationship between RNA polymerase and the Supercoiled template precludes circling of the RNA polymerase around the DNA and prevents the growing RNA transcript from becoming entangled with the template DNA.
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direct visualization of Supercoiled DNA molecules in solution
The EMBO Journal, 1990Co-Authors: Marc Adrian, Ten B Heggelerbordier, Walter Wahli, Alicja Z Stasiak, Jacques DubochetAbstract:The shape of Supercoiled DNA molecules in solution is directly visualized by cryo-electron microscopy of vitrified samples. We observe that: (i) Supercoiled DNA molecules in solution adopt an interwound rather than a toroidal form, (ii) the diameter of the interwound superhelix changes from about 12 nm to 4 nm upon addition of magnesium salt to the solution and (iii) the partition of the linking deficit between twist and writhe can be quantitatively determined for individual molecules.
