The stretched intermediate model of B-Z DNA transition

There have been numerous attempts to describe the mechanism of B-Z transition. Our simulations based on the stochastic difference equation with length algorithm show that a short DNA oligomer will tend to unwind and overstretch during the transition. The overstretching of DNA is then understood from the Zhou, Zhang, and Ou-Yang model. Unlike the Harvey model, the stretched intermediate model does not pose any steric dilemma; we are able to show that the chain sense reversal progresses spontaneously using the stretched intermediate model. A nonlinear DNA model is used to describe the origins and mechanism of base rotation in the stretched intermediate state of DNA. We also propose an experiment that can verify the existence of a stretched intermediate state during B-Z transition, thus opening up fresh grounds for experimentation in this field.     

B-A and B-Z transitions in deoxyoligoduplexes containing 4- and 5- methylcytosine

Self-complementary oligodeoxynucleotides: GGACCCGGGTCC, GGA4mCCCGGGTCC, GGA5mCCCGGGTCC, CGCGCGCG, CG4mCGCGCG, CG5mCGCGCG were synthetized to study the contribution of methyl groups into the energetics of the three known cooperative transitions in DNA: helix-coil, B-A and B-Z With the use of circular dichroism and absorbtion methods the profiles of the above transitions were obtained by variation of temperature (helix-coil), trifluoroethanol fraction (B-A), NaCl and trifluorethanol contents (B-Z). On the basis of the transition widths and shifts of the transition points due to the methylations the energetics of the methyl groups was estimated. 5mC stabilizes the B form relatively the A form by 0.33 kcal/mol; while 4mC by 0.5 kcal/mol. In the B-Z transition 5 mC stabilizes the Z form by 0.28 kcal/mol relatively the B form; 4mC stabilizes also the Z form although by 0.14 kcal/mol only. Thus, these naturally occurring modifications could modulate substantially the ability of a DNA piece to shift into the A or Z form.     

Salt dependence of DNA structural stabilities in solution. Theoretical predictions versus experiments

The predictions of currently available theories for treating DNA-diffuse ionic cloud free energy contributions to conformational stability have been tested against experimental data for salt induced B-Z and B-A transitions. The theories considered are (i) Manning's counterion condensation approach (CC), (ii) the idealized Poisson-Boltzmann approximation (PB), and (iii) the potentials of mean force (PMF) approach proposed by Soumpasis. As far as we can judge from comparison with the set of experimental data currently available, it is found that only the latter theory yields satisfactory quantitative results for the dependence of the B-Z and B-A relative stabilities on monovalent salt concentration. The correct application of the PB and CC theories does not yield very low salt Z-B transitions, in contradiction to earlier assertions. At low salt concentrations the PB theory is qualitatively correct in predicting that the B form is electrostatically more favorable than both the A and Z forms, whereas the CC theory is qualitatively wrong predicting that Z-DNA is more stable than both B and A DNA.     

The energetics of the B-Z transition in DNA

The paper deals with the energetics of the transition to left-handed Z form in DNA with an arbitrary base sequence. There is a brief outline of the statistical-mechanical model of the B-Z transition allowing for three possible states of each base pair. The parameters of the model can be determined by comparing the theory with experimental data for the B-Z transition in inserts with given sequences in circular DNA. The model contains six energy parameters, most of which have been determined before. In order to find the remaining parameters of the model and test its adequacy, a number of oligonucleotide sequences were synthesized and inserted into the pUC 19 plasmid. Two-dimensional gel electrophoresis was used to determine the superhelical density at which the inserts adopt the Z form. A statistical-mechanical treatment of these data yielded a complete set of six energy parameters for the B-Z transition. The theoretical assumption that the free energy of Z-form pairs does not depend on the type of adjacent pairs proved to be in agreement with the experimental data.     

The Energetics of the B-Z Transition in DNA

Abstract

The paper deals with the energetics of the transition to left-handed Z form in DNA with an arbitrary base sequence. There is a brief outline of the statistical-mechanical model of the B-Z transition allowing for three possible states of each base pair. The parameters of the model can be determined by comparing the theory with experimental data for the B-Z transition in inserts with given sequences in circular DNA The model contains six energy parameters, most of which have been determined before. In order to find the remaining parameters of the model and test its adequacy, a number of oligonucleotide sequences were synthesized and inserted into the pUC 19 plasmid. Two-dimensional gel electrophoresis was used to determine the superhelical density at which the inserts adopt the Z form. A statistical-mechanical treatment of these data yielded a complete set of six energy parameters for the B-Z transition. The theoretical assumption that the free energy of Z-form pairs does not depend on the type of adjacent pairs proved to be in agreement with the experimental data.     

Energetics of B-Z junction formation in a sixteen base-pair duplex DNA

We report analysis of the NaCl-induced B-Z transition in a 16 base-pair duplex DNA with sequence designed such that when NaCl is increased the left half of the molecule undergoes the B-Z transition while the right half remains in the B-form. An equilibrium thermodynamic model based on the body of available published experimental data and the recent theoretical work of Soumpasis, which indicate, in the salt range above approximately 0.9 M-NaCl, the transition free-energy of B-Z conversion in DNA is a linear function of the NaCl concentration, is employed. Analysis of the B-Z transition of the junction-containing molecule indicates the B-Z junction formed in this 16 base-pair DNA is composed of approximately three base-pairs and has a free energy of formation of approximately 4.7 kcal/mol junction. These values for the junction are in excellent agreement with published estimates of B-Z junction size and energy derived from much longer DNA pieces.     

Salt Dependence of DNA Structural Stabilities in Solution. Theoretical Predictions Versus Experiments

Abstract

The predictions of currently available theories for treating DNA-diffuse ionic cloud free energy contributions to conformational stability have been tested against experimental data for salt induced B-Z and B-A transitions. The theories considered are (i) Manning's counterion condensation approach (CC), (ii) the idealized Poisson-Boltzmann approximation (PB), and (iii) the potentials of mean force (PMF) approach proposed by Soumpasis. As far as we can judge from comparison with the set of experimental data currently available, it is found that only the latter theory yields satisfactory quantitative results for the dependence of the B-Z and B-A relative stabilities on monovalent salt concentration. The correct application of the PB and CC theories does not yield very low salt Z-B transitions, in contradiction to earlier assertions. At low salt concentrations the PB theory is qualitatively correct in predicting that the B form is electrostatically more favorable than both the A and B forms, whereas the CC theory is qualitatively wrong predicting that Z-DNA is more stable than both B and A DNA.     

The influence of tertiary structural restraints on conformational transitions in superhelical DNA.

This paper examines theoretically the effects that restraints on the tertiary structure of a superhelical DNA domain exert on the energetics of linking and the onset of conformational transitions. The most important tertiary constraint arises from the nucleosomal winding of genomic DNA in vivo. Conformational transitions are shown to occur at equilibrium at less extreme superhelicities in DNA whose tertiary structure is restrained than in unrestrained molecules where the residual linking difference alpha res (that part of the superhelical deformation which is not absorbed by transitions) may be freely partitioned between twisting and bending. In the extreme case of a rigidly held tertiary structure, this analysis predicts that the B-Z transition will occur at roughly half the superhelix density needed to drive the same transition in solution, other factors remaining fixed. This suggests that superhelical transitions may occur at more moderate superhelical deformations in vivo than in solution. The influence on transition behavior of the tertiary structural restraints imposed by gel conditions also are discussed.     

Left-handed Z form in superhelical DNA: a theoretical study

This is a comprehensive statistical mechanical treatment of the Z form formation in purinepyrimidine stretches of different length inserted into superhelical DNA. The B-Z transition for short inserts is shown to follow the "all-or-none" principle. Over some critical value of the insert length n, the B-Z transition in the insert proceeds in two stages. The flipping of m base pairs into the Z form is followed by a gradual growth of the Z-form stretch until it occupies the whole insert. By fitting the theoretical transition curves to experimental ones the fundamental thermodynamic parameters of the B-Z transition have been determined: the B-Z junction energy Fj = 4-5kcal.mol-1 and the free energy change delta FBZ = 0.5-7.0 kcal.mol-1 under standard salt conditions. Calculations show that the B-Z transition in short purinepyrimidine inserts may be seriously affected by cruciform formation in the carrier DNA.     

Left-handed Z Form in Superhelical DNA: A Theoretical Study

Abstract

This is a comprehensive statistical mechanical treatment of the Z form formation in purine- pyrimidine stretches of different length inserted into superhelical DNA. The B-Z transition for short inserts is shown to follow the “all-or-none” principle. Over some critical value of the insert length n, the B-Z transition in the insert proceeds in two stages. The flipping of m base pairs into the Z form is followed by a gradual growth of the Z-form stretch until it occupies the whole insert. By fitting the theoretical transition curves to experimental ones the fundamental thermodynamic parameters of the B-Z transition have been determined: the B-Z junction energy Fj = 4–5kcal?mol^?1 and the free energy change ΔF_B-Z = 0.5–0.7 kcal?mol^?1 under standard salt conditions. Calculations show that the B-Z transition in short purine-pyrimidine inserts may be seriously affected by cruciform formation in the carrier DNA.     

AFM studies of DNA structures on mica in the presence of alkaline earth metal ions

As counterions of DNA on mica, Mg(2+), Ca(2+), Sr(2+) and Ba(2+) were used for clarifying whether DNA molecules equilibrate or are trapped on mica surface. End to end distance and contour lengths were determined from statistical analysis of AFM data. It was revealed that DNA molecules can equilibrate on mica when Mg(2+), Ca(2+) and Sr(2+) are counterions. When Ba(2+) is present, significantly crossovered DNA molecules indicate that it is most difficult for DNA to equilibrate on mica and the trapping degree is different under different preparation conditions. In the presence of ethanol, using AFM we have also observed the dependence of B-A conformational transition on counterion identities. The four alkaline earth metal ions cause the B-A transition in different degrees, in which Sr(2+) induces the greatest structural transition.     

A study of the B-A transition in DNA by gel electrophoresis

A procedure is developed for studying the B-A transition in DNA using gel electrophoresis. The starting point has been the idea that the junction between the A and B sections, which appear within the transition interval would increase the mobility of the DNA molecules. Indeed, the mobility of DNA in a gel is shown to increase in the middle of the B-A transition due to the formation of the largest possible number of boundaries between the B and A forms. The middle of the B-A transition in supercoiled DNA appears to be shifted against the middle of the transition in open circular (as well as linear) DNA by about 1.3% towards lower ethanol concentrations under the influence of the superhelical stress.     

Energetics of superhelicity and of B-Z transitions in superhelical DNA

The linking difference, alpha, imposed upon a superhelically constrained DNA molecule must be partitioned between twisting and bending deformations. Transitions to alternative secondary structures can occur at susceptible sites, altering the local molecular twist by an amount delta Twtrans. That part of the linking difference not accommodated in this way, the residual linking difference alpha res, must be manifested as smooth torsional and flexural deformations of secondary structure. The competition among the alternative ways of accommodating the imposed linking difference alpha determines a stressed equilibrium state. The superhelical free energy, G(alpha), is the excess free energy of the equilibrium state at linking difference alpha above that of the relaxed state under identical conditions. In this paper a method is described by which the free energies associated both to linking, G(alpha), and to residual linking differences can be determined from data on superhelical conformational transitions. The application of this approach to previously published experimental data on the B-Z transition suggests that the free energy associated with alpha res is about 30% larger at substantial superhelicities than it is near the relaxed state. At the onset of transition the functional form of G(alpha) is shown to change in a manner dependent upon the length of the Z-susceptible site.     

A Study of the B-A Transition in DNA by Gel Electrophoresis

Abstract

A procedure is developed for studying the B-A transition in DNA using gel electrophoresis. The starting point has been the idea that the junction between the A and B sections, which appear within the transition interval would increase the mobility of the DNA molecules. Indeed, the mobility of DNA in a gel is shown to increase in the middle of the B-A transition due to the formation of the largest possible number of boundaries between the B and A forms. The middle of the B-A transition in supercoiled DNA appears to be shifted against the middle of the transition in open circular (as well as linear) DNA by about 1.3% towards lower ethanol concentrations under the influence of the superhelical stress.     

A model of the strain-induced B-Z transition

The B-to-Z transition in supercoiled circular DNA is modeled as a strain-induced nonlinear excitation process. Using a model, in which DNA is regarded as a chain of units with a bistable energy function along the twisting coordinate together with a harmonic inter-unit interaction, we show that a Z region and the accompanying two B-Z junctions of finite width appear naturally as a solution of nonlinear equations, when the strain exceeds a critical value. We examine the B-Z transition behaviour as a function of twist under various situations. We also analyse available experimental results on B-Z transition in supercoiled plasmid with G-C insertions by this mechanistic model in order to estimate the magnitude of model parameters. The energy barrier of the B-Z transition is estimated to be of the order of 1 kcal/mole per base pair. The analysis shows that if the length of the insertion is less than a certain value, the entire insertion converts to Z form at a transition point, but if the insertion is much longer, the B-Z transition exhibits a different behavior, in which part of the insertion flips to Z form and the Z region expands linearly upon changing linking number.     

Mapping the B-A conformational transition along plasmid DNA

A simple method is presented to monitor conformational isomerizations along genomic DNA. We illustrate properties of the method with the B-A conformational transition induced by ethanol in linearized pUC19 plasmid DNA. At various ethanol concentrations, the DNA was irradiated with ultraviolet light, transferred to a restriction endonuclease buffer and the irradiated DNA was cleaved by 17 restriction endonucleases. The irradiation damaged DNA and the damage blocked the restrictase cleavage. The amount of uncleaved, i.e. damaged, DNA depended on the concentration of ethanol in a characteristic S-shape way typical of the cooperative B-A transition. The transition beginning and midpoint were determined for each restriction endonuclease. These data map the B-A transition along the whole polylinker of pUC19 DNA and six evenly distributed recognition sequences within the rest of the plasmid. The transition midpoints fell within the B-A transition region of the plasmid simultaneously determined by CD spectroscopy. The present method complements the previous methods used to study the B-A transition. It can be employed to analyze multikilobase regions of genomic DNA whose restriction endonuclease cleavage fragments can be separated and quantified on agarose gels.