Helix-coil Transition Region Research Articles

Near-neighbor effects in cooperative modified self-consistent phonon approximation melting in DNA.

The modified self-consistent phonon approximation approach is a microscopic theory describing DNA thermal fluctuational motion both in the premelting temperature regime and in the helix-coil transition region. In the present paper we show that the near-neighbor effects can be introduced into the theory by a simple extension in the logic of the cooperativity. The present theory can give base-sequence effects on melting as well as premelting behavior in agreement with experiments. These effects arise from the same interatomic interaction model used in our earlier calculations [Biopolymers 33, 351 (1993); 33, 797 (1993)] and the results do not require the explicit introduction of near-neighbor parameters. The work then becomes a no-parameter derivation of near-neighbor effects in DNA.

A cooperative self‐consistent microscopic theory of thermally induced melting of a repeat sequence DNA polymer

AbstractWe attempt to extend the modified self‐consistent phonon theory to describe thermal fluctuational base‐pair opening of repeat sequence DNA polymers in the helix–coil transition region as well as in the premelting region. A microscopic base‐pair open state is introduced and the effect of this open state is taken into account self‐consistently in a mean field system that models the DNA polymer. Our analysis indicates the structure of this open state changes with temperature in such a manner that on average a base pair opens and unstacks with its neighbors more completely as temperature increases. We apply this theory to a homopolymer—poly(dG) · poly(dC) to evaluate the base‐pair opening probability in a temperature range from 273 to 366.5 K. At 366.5 K the system undergoes cooperative melting. Our calculated base‐pair opening probabilities are in general agreement with several experimental estimates at room temperature. The calculated probabilities show typical melting curve behavior at temperatures close to the observed melting temperature. The cooperative modified self‐consistent phonon approximation approach becomes a viable microscopic theory of melting. © 1993 John Wiley & Sons, Inc.

Theory of pressure-dependent melting of the DNA double helix: Role of strained hydrogen bonds.

In this paper we study the effect of hydrostatic pressure on the thermal stability of DNA base pairs. The thermal stability of a base pair is predominantly determined by the fluctuational motion of the interbase hydrogen bonds in the base pair. We postulate that the pressure exerted on the surface of DNA creates a compressive stress in the interbase hydrogen bonds. This stress in turn induces a strain in these hydrogen bonds. The effect of this compressive stress on the thermal fluctuational motion of the strained interbase hydrogen bonds in DNA can be analyzed by the modified self-consistent phonon approximation theory. We calculate individual hydrogen bond disruption probabilities and base pair opening probabilities of two B-conformation DNA polymers, an adenine-thymine copolymer and a guanine-cytosine homopolymer, at various pressures and at temperatures both in the premelting region and in the helix-coil transition region. Our calculated pressure dependence of the melting temperature for both polymers is in fair agreement with experimental observations. Our values for the melting temperatures are also in agreement with observation and our theory is successful as a theory of cooperative melting.

Premelting thermal fluctuational base pair opening probability of poly(dA).poly(dT) as predicted by the modified self-consistent phonon theory.

We employ a mean field, modified, self-consistent phonon theory to evaluate the single base-pair opening rate and the probability of a base pair in the amino proton exchangeable state for the homopolymer poly(dA).poly(dT) at temperatures below the helix-coil transition region. Our calculated premelting single base-pair opening probabilities are in general agreement with several available experimental estimates from imino proton exchange and formaldehyde-induced DNA melting measurements. These calculated opening probabilities, however, are in disagreement with the prediction of the helix-coil transition theory. Possible reasons for the differences are discussed, especially the possible different definition of a meaningful open state in the premelting region. The premelting open state of the modified self-consistent phonon approximation theory seems to be appropriate to describe a solvent-accessible open configuration that is sufficient to facilitate important chemical reactions such as imino proton exchange and formaldehyde reaction with the bases. This can be compared with the completely unstacked open state of the helix-coil transition theory originally defined in the helix-coil transition region. We propose that the amino proton exchangeable state is different from the open state associated with melting and only involves the breaking of the amino interbase H bond. The agreement between the calculated and experimentally estimated probability of a base pair in the amino proton exchangeable state seems to support this hypothesis.

Non‐newtonian viscosity of polypeptides in the helix–coil transition region

AbstractThe non‐Newtonian intrinsic viscosities [η] of poly(γ‐methyl L‐glutamate) were measured in the helix–coil transition region under various conditions in this work. The helix content fH, which represents the degree of conformational transition, was obtained by using a polarimeter. Our experimental results show that the non‐Newtonian behavior of the polypeptide is markedly affected by its conformation; i.e., the non‐Newtonian effect becomes larger as fH increases. The effect of external pressure ΔP on [η] was studied carefully; [η] increases with fH when ΔP < 1.5 psi, but it decreases when ΔP > 1.5 psi and fH > 0.8. The reason for this result is considered in the text.

Deuteration kinetics of deoxyguanosine, deoxycytidine, and their polynucleotides by means of ultraviolet spectrophotometry.

The kinetics of the hydrogen-deuterium exchange reactions of deoxyguanosine (dG), deoxycytidine (dC), double-helical poly[d(G-C)] X poly[d(G-C], and double-helical poly(dG) X poly(dC) have been examined at 20 degrees C, pH 7.0, and in low-salt (0.15 M NaCl) medium by stopped-flow ultraviolet spectrophotometry, in the spectral region of 260 to 320 nm. The rate constant was found to be 78.9 s-1 for dG-NH, 2.2 s-1 for dG-NH2, 39.3 s-1 for dC-NH2, 2.4 s-1 (fast) and 0.94 s-1 (slow) for poly[d(G-C)] X poly[d(G-C)], and 2.2 s-1 (fast) and 0.92 s-1 (slow) for poly(dG) X poly(dC). From these values, the probability of base-pair opening of the G X C containing B-form double helix is estimated to be (3 +/- 1) X 10(-3). This is much greater than what is expected from an extrapolation of the van't Hoff plot at the helix-coil transition region, i.e. at about 110 degrees C. The mechanism of these base-pair openings at 20 degrees C (as well as the mechanism of base-pair reformation) is suggested to be totally different from those in the melting temperature range.

Dimensional Properties of Polypeptides in the Helix–Coil Transition Region II. Intrinsic Viscosity

The theory presented in the preceidng paper (Part I) for the molecular dimensions, ‹s2›oI and ‹s2›I, of polypeptides in the helix–coil transition region is applied here to determine the intrinsic viscosity [η]I, (the [η] in the transition region) as a function of the helical content fH which varies with the cooperativity parameter (eq I-16). Using the Kurata–Yamakawa theory, the linear expansion factor αs was converted to the hydrodynamic expansion factor αη Thus, with αηI, (the αη in the transition region) and ‹s2›oI (eq I-10), the [η]I’s of polypeptides in the helix–coil transition region were calculated (eq 3), and compared with experimental results for various kinds of polypeptide–solvent systems with good agreement (Figures 3 and 4).

Dimensional Properties of Polypeptides in the Helix–Coil Transition Region I. Molecular Dimension

An expression was derived to correlate the unperturbed mean-square radius of gyration of an intermediate polypeptide chain ‹S2›oI with helical content fH (see eq 10). An unperturbed state corresponding to the α-helix was obtained by converting the helix to a tortuous random-coil with a bond-rotation angle of +100° or −100° (in cis-convention) per residue, or per “virtual bond,” and a bond angle satisfying the experimental value of 8.8 for the characteristic ratio of the random-coil of PBLG. The “quasi” linear expansion factor αh of a helical sequence in an intermediate chain was evaluated (see eq 18) using the Zimm–Bragg–Nagai theory in collaboration with our expression for ‹S2›o. The intermediate chain may be a kind of swollen random-coil owing to the existence of helical sequences in the chain, and thus, the linear expansion factor αsI of the entire intermediate chain was approximately formulated as αsI=αsCαh where αsC is the linear expansion factor for the radius of gyration of a random-coil. With αsI and ‹S2›oI, the mean-square radius of gyration ‹S2›I of polypeptides in the transition region was calculated (eq 12) and compared with that obtained experiment (see Figure 4). Our theoretical results were found to agree well with experiment.

Scattering functions of poly(amino acid) in helix-coil transition

Scattering functions of poly(amino acid) in the helix-coil transition region have been calculated on the basis of the Zimm-Bragg theory, to examine the effects of the helical content on the scattering function and on the cross-section factor, from which the radius of gyration of the cross section and the mass per unit length of the helical sequence can be obtained. The molecule is assumed to contain only one α-helical sequence of any length, leaving unfolded coils at the chain ends. The statistical mechanical averages of the interference among scattering units, apart from the interference within the helical sequence, are approximated by the expansion in terms of the even moments up to the eighth moment, which are calculated by the generator matrix method based on the conformation energies computed for poly ( l-alanine). The scattering function at intermediate angles varies systematically according to the variation of the helical content, demonstrating the sensitivity of intermediate-angle scattering to the conformational changes in the helix-coil transition. The comparison of the Guinier plots of the cross-section factors, calculated from the scattering functions at various helical contents, has indicated that the radius of gyration of the cross section of the helical sequence can be determined accurately from the plots even when the coil content is as much as 30%, and that the mass per unit length is determinable to within an error of 6% unless the coil content exceeds 10%.

Fluctuational base‐pair opening in DNA at temperatures below the helix–coil transition region

Fluctuational base‐pair opening in DNA at temperatures below the helix–coil transition region

Light scattering from poly(L-glutamic acid) in aqueous solution in the helix-coil transition region

L'interaction electrostatique due aux charges sur les chaines laterales influence les dimensions moleculaires du polyaminoacide dans la region de la transition helice pelote

Fluctuational base-pair opening in DNA at temperatures below the helix-coil transition region.

AbstractThe probability of DNA base‐pair opening was calculated at temperatures below the denaturation region. The helix–coil transition theory, modified to include different nearest‐neighbor interactions, was employed. Predictions of base‐pair opening employing DNA melting‐curve parameters differed considerably from predictions based on parameters evaluated from synthetic RNA oligomer data of Gralla and Crothers (G‐C) and formaldehyde–DNA binding experiments of McGhee and von Hippel (M‐vH). Calculations based on the latter parameters indicate a base‐pair‐opening probability of 10−2–10−3 at 35°C in 0.1M NaCl. DNA melting‐curve parameters predict values about 103 smaller. At a temperature 10°C below the transition midpoint of a specific DNA sequence, DNA melting parameters predict base‐pair opening of about 10−3, whereas the G‐C and M‐vH parameters predict that ≳0.16 of the DNA is melted. Experiments and theoretical assumptions relevant to the calculations are analyzed. Evidence suggests that DNA melting parameters are valid when the average loop size exceeds some minimum value, whereas the G‐C and M‐vH parameters appear more valid for single base‐pair loops. A reconciliation of the two sets of predictions can be made if interactions extending beyond neighboring base pairs are considered. Such interactions will make the parameters of the nearest‐neighbor model appear to change with the average loop size. Experiments that may provide further measurements of base‐pair opening are discussed.

The helix-coil transition of dna restriction fragments containing the lactose operon promoter sequence

Abstract The theory of the duplex to single stranded transition of DNA has been employed to calculate differential melting curves for DNA fragments, 95, 144, 203 and 301 base pairs (bp.) in length. The base pair sequences of these DNAs are known (Dickson et. al. Science 182, 65, 1977). These DNAs contain the 40–50 bp. promoter sequence which is bound by RNA polymerase to initiate transcription. Comparison was made to experimental melting curves of the transitions in 0.1M Na+ solution. The theory included the effects of DNA length on final dissociation. Values of theoretical parameters were obtained from previous theory-experiment comparisons of natural and synthetic DNAs. Except for β, the dissociation constant for final dissociation to single strands a single set of parameters was used. As expected β varied with the length of the fragment. Theoretical calculations produced agreement with the fine structure and Tm values of experimental curves which was within experimental error. Melting profiles calculated at temperatures within the transition inducated the DNAs melt from the ends with blocks of instability from 10 to 60 bp. long. The theory was also employed to examine the probability of base pair opening within the lactose promoter at temperatures below the helix-coil transition region. At 37°C the fluctuational opening of base pairs depends very little on its sequence environment. The average probability of opening, Θ, is 3.5×10−6 for A·T bps. and 6.9×10−7 for G·C bps. As one approaches the transition, the influence of the sequence environment increases. At 77°C, the average Θ values increase by 10 fold from 37°C. However the range of Θ for A·T pairs varies up to 200% compared to 6% for the same sequence at 37°C.

An Application of the Extended Principal Component Analysis to Circular Dichroism and Optical Rotatory Dispersion. A Case of Poly(α-l-glutamic acid) in the Helix-Coil Transition Region

Abstract The extended principal-component-analysis (PCA) method has been applied to circular dichroism (CD) and optical rotatory dispersion (ORD). The validity of this method has been examined with ideal binary and ternary equilibrium systems, for which the data CD spectra and ORD curves were computer-generated. Satisfactory results were obtained for these multicomponent optically active systems. As an example, the ultraviolet CD spectra of poly(α-l-glutamic acid) in the pH-induced helix-coil transition region were analyzed with the PCA procedure. By the extended PCA method, the pure CD spectra for both helix and random-coil conformations and the helix fractions in the transition region were determined from eight experimentally obtained CD spectra. By introducing an empirical parameter α into the Henderson-Hasselbalch equation, the helix-coil transition curve was reproduced reasonably well.

Spectroscopic and equilibrium dialysis studies of poly(α‐L‐glutamic acid)–Cu(II) complexes in the pH range 4–7

AbstractThe formation of complex between the Cu2+ ion and poly(α‐L‐glutamic acid) [poly(Glu)] in 150 mM NaCl solutions was studied by uv–visible absorption and equilibrium dialysis methods at the mixing ratios of Glu residues to Cu2+, R, of 32, 16, and 8 and in the pH range 4–7. The results showed that more than 90% of Cu2+ ions bind to the poly(Glu) at pH > 4.9, but the bound Cu(II) begins to dissociate with a decrease in pH. The absorption spectra of bound Cu(II) varied with pH and R in a complicated manner. Three different component spectra were disclosed from the analysis of the pH dependence of the bound spectra. We concluded that poly(Glu)–Cu(II) complexes fall into three classes in the pH range 4–7, with the proportions of these complexes varying with both pH and R. The three complexes predominate either in the helix or extended‐coil region, in the helix–coil transition region, or in the helix‐aggregate region. The stability constant and binding mode of each Cu(II)–Glu complex were estimated from the dialysis data. With these results, the possible structure of each complex is discussed.

Equilibrium Dialysis Study of the Interaction between Poly(α-L-glutamic acid) and Transition Metal Ions in the Helix–Coil Transition Region

By the combined equilibrium dialysis-colorimetric method, the fractions, fb, of bound Ni(II) and Cd(II) in the poly(α-L-glutamic acid)–Ni(II) and–Cd(II) solutions were determined at 25°C over a pH range of 7—4.2. Both Ni2+ and Cd2+ ions were dissociated by about 25% at pH 6.5 (fb≍75%), where poly(Glu) is in the random–coil form, and by nearly 90% at pH 4.5, where poly(Glu) is almost a complete helix. The ORD data of poly(Glu)–Ni(II) and–Cd(II) complexes were essentially the same as those of poly(Glu), indicating that these transition metal ions do not induce the helical structure in poly(Glu). This is a striking contrast with the case of Cu2+. From the equilibrium dialysis data, it was concluded that each Ni2+ or Cd2+ ion binds with two adjacent glutamyl residues in poly(Glu)–Ni(II) and –Cd(II) complexes. The stability constants of these complexes were estimated to be of the order of 103.

Origin of multipeak behavior in the NMR spectra of poly(L‐glutamic acid)

AbstractThe previous report that poly(L‐glutamic acid) exhibits doubled resonances in the helix–coil transition region by either proton or carbon‐13 nmr resolves the question of whether or not this behavior is limited to uncharged polypeptides in organic solvents, as had been previously thought. In the present work, we show that the underlying principle causing this anomalous double‐peak behavior is due to molecular‐weight polydispersity of the sample. The molecular‐weight range in which this phenomenon is observed is largely dependent on the values of σ, the nucleation or cooperativity factor. The principles developed are shown to encompass all classes of polypeptides in a very natural way and to explain the key experimental data in the literature.

Conformational Lifetimes of the Helix—Coil Transition of Polypeptides

In order to explain the separate α-CH peaks in NMR spectra of polypeptides in the helix—coil transition region, the lifetimes of molecules in two sites (one being a totally random-coil conformer and the other, comprising all other conformers containing at least one helix unit), were calculated on the basis of the one-dimensional Ising model in accordance with a two-site model. A lifetime greater than 10−2 s was not obtainable for a totally random-coil conformer for chain lengths ranging from 10 to 85. It was concluded that the occurrence of the separate peaks could not be attributed to the slow exchange rate between the two sites. Polydispersity in the chain length seems to be major consequence of such separate peaks.

A nuclear magnetic resonance study of the helix-coil transition of poly(L-glutamic acid).

AbstractThere have been many reports that the nuclear magnetic resonance (nmr) spectra of a large number of polypeptides exhibit peak doubling of the α‐carbon and the α‐carbon proton in the helix–coil transition region. One apparent exception to this generalization has been polypeptides with ionizable side chains, where the helix–coil transition is induced by changes in pH in aqueous solution. Because it is important to establish the proper theoretical reason for the peak doubling and its relation to the rate of conformational change of amino acid residues, we have reexamined the proton and carbon‐13 nmr spectra, at high field, for two polydisperse samples of poly(L‐glutamic acid). Doubling of the α‐carbon proton resonance as well as those of the α‐ and β‐carbon, and backbone carbonyl are observed for a low‐molecular‐weight sample (DP = 54), while a higher molecular weight sample (DP = 309), exhibits only single resonances. Thus, polydispersity by itself is not sufficient to observe peak doubling; low‐molecular weight is also required.

Intensity fluctuation spectroscopy of deoxyribonucleic acid (DNA). II. Temperature profile of bacteriophage phi29-DNA.

AbstractIntensity‐fluctuation laser‐scattering spectroscopy measurements have been reported for the DNA isolated from bacteriophage ϕ29. The cumulant method of data analysis gives well‐defined weighted average decay frequencies. Available theoretical models are shown to yield good values for the diffusion coefficient and longest intramolecular relaxation time for low‐molecular‐weight DNA's but not for higher molecular weight species. The temperature dependence of the apparent diffusion coefficient exhibits drastic oscillations in the helix–coil transition region. The number of these oscillations is consistent with independent measurements of uv hyperchromicity measurements and suggests that ϕ29‐DNA is composed of at least three regions of differing G + C content.