Abstracts of 1996 Publications


Rouzina, I. and Bloomfield, V.A. (1996) Competitive Electrostatic Binding of Charged Ligands to Polyelectrolytes: Planar and Cylindrical Geometries. J. Phys. Chem. 100: 4292-4304.

The strong salt dependence of the binding of multivalent ligands to DNA indicates the predominantly electrostatic nature of that binding. To a good first approximation, the salt dependence can be expressed as a competition between two point counterion species in screening the highly charged macroion surface. This basic problem is solved in terms of the Poisson-Boltzmann (P-B) equation for a macroion of arbitrary shape and charge, in a solution of arbitrary salt composition and ionic strength. Various binding regimes are defined by the relative magnitudes of a, the radius of curvature of the macroion; rd, the Debye-Huckel screening length, which characterizes exponential decay in the linear screening regime; and lambda, the decay length of the counterion distribution close to the macroion surface in the nonlinear screening regime. Only the nonlinear regime (lambda « rd) produces relatively high free energy of ligand binding (> kBT per ion). This regime is very similar in planar and cylindrical geometries. For this case we suggest a simple new method of calculating the amount of each species bound, which avoids numerical solution of the P-B equation for each set of species concentrations and further integration of the charge. Instead, we follow changes in the surface concentration and the decay length of each counterion distribution in the course of titration. The product of these two quantities yields a reasonably accurate estimate for the amounts bound. It also yields a closed form of the binding isotherm in both geometries. The apparent ligand (species 2) binding constant obtained in this way has a conventional dependence on the salt (species 1) concentration to the -z2/z1 power. However, we obtain a new expression for the magnitude of the electrostatic binding constant in terms of the macroion surface charge density, Bjerrum length, and ion charges z1 and z2. Binding of counterions to the DNA double helix changes qualitatively from nonlinear cylindrical, through nonlinear planar, to linear planar behavior as the ionic strength of the solution is raised. Our results are comparable in many ways to those of counterion condensation theory and the McGhee-von Hippel site binding model, but show some different behavior and suggest different interpretations of similar behavior.


Rouzina, I. and Bloomfield, V.A. (1996) Influence of ligand spatial organization on competitive electrostatic binding to DNA. J. Phys. Chem. 100: 4305-4313.

The influence of ligand size on electrostatic binding to DNA in a background of competing counterions is analyzed with the planar Poisson Boltzman (P-B) equation, which is analytically integrated in two screening layers near the polyion surface: one sterically accessible only to the smaller counterions, and the other accessible to both species. We obtain an explicit expression for the dependence of the electrostatic binding constant on the difference in counterion radii d. This dependence is approximately exponential with decay length lambda/z2, where lambda is the thickness of the screening layer due to the smaller species and z2 is the ligand valence. lambda is determined by the surface charge density of the polyion and the Bjerrum length, and is a few Å for double-helical DNA. This approach agrees well with detailed P-B and Monte Carlo calculations for mixtures of hard sphere counterions, and accounts for NMR results on the competitive binding of monovalent and divalent cations. The distribution of charges on the ligand is shown to affect the power S through which the salt concentration enters the ligand binding constant. If the counterion species have similar radii (Dd < lambda), their competition is well described by a point charge model, which gives the conventional S = z2/z1. If the ligand is much larger than the salt counterions (Dd » lambda), it does not participate in screening at all, so S = 0. An intermediate difference in counterion radii (Dd l) results in an effective ligand charge zeff different from z2. If the ligand is larger than the salt counterion, then zeff < z2. The opposite situation is also possible depending on ligand structure. We propose a simple method for the approximate prediction of the effective charge in competitive electrostatic binding of a ligand with distributed charges.


Bloomfield, V.A. (1996) DNA condensation. Curr. Opinion Struct. Biol. 6: 334-341.

Recent progress in our understanding of DNA condensation includes observation of the collapse of single molecules, greater insight into the intermolecular forces driving condensation, recognition of helix structure perturbation in condensed DNA, and increasing recognition of the likely biological consequences. DNA condensed with cationic liposomes is an efficient agent for transfection of eukaryotic cells, with considerable potential interest for gene therapy.


Duguid, J.G., and Bloomfield, V.A. (1996) Electrostatic effects on the stability of condensed DNA in the presence of divalent cations. Biophys. J. 70: 2838-2846.

Cylindrical cell model Poisson-Boltzmann (P-B) calculations are used to evaluate the electrostatic contributions to the relative stability of various DNA conformations (A, B, C, Z, and single-stranded (ss) with charge spacings of 3.38 and 4.2 A) as a function of interhelix distance in a concentrated solution of divalent cations. The divalent ion concentration was set at 100 mM, to compare with our earlier reports of spectroscopic and calorimetric experiments which demonstrate substantial disruption of B-DNA geometry. Monovalent cations which neutralize the DNA phosphates in two ways, corresponding to different experimental situations: (1) There is no significant contribution to the ionic strength from the neutralizing cations, corresponding to DNA condensation from dilute solution, and to osmotic stress experiments in which DNA segments are brought into close proximity in the presence of a large excess of buffer. (2) The solution is uniformly concentrated in DNA, so that the neutralizing cations add significantly to those in the buffer at close DNA packing. In case (1), conformations with lower charge density (Z and ssDNA) have markedly lower electrostatic free energies than B-DNA as the DNA molecules approach closely, due largely to ionic entropy. If the divalent cations bind preferentially to single stranded DNA or a distorted form of B-DNA, as is the case with transition metals, the base pairing and stacking free energies which stabilize the double helix against electrostatic denaturation may be overcome. Strong binding to the bases is favored by the high concentration of divalent cations at the DNA surface arising from the large negative surface potential; the surface concentration increases sharply as the interhelical distance decreases. In case (2), the concentration of neutralizing monovalent cations becomes very large and the electrostatic free energy difference between secondary structures becomes small as the interhelical spacing decreases. Such high ionic concentrations will be expected to modify the stability of DNA by changing water activity as well as by screening electrostatic interactions. This may be the root of the decreased thermal stability of DNA in the presence of high concentrations of magnesium ions.


Rouzina, I. and Bloomfield, V.A. (1996) Macroion Attraction Due to Electrostatic Correlation Between Screening Counterions. I. Mobile Surface-Adsorbed Ions and Diffuse Ion Cloud. J. Phys. Chem. 100: 9977-9989.

Highly charged macroion surfaces in solutions of multivalent electrolytes attract each other electrostatically through correlations in their counterion environment. We show that significant correlation occurs when the counterion distribution has a pseudo two-dimensional character. This allows us to treat the electrostatic correlation attraction semianalytically by reducing the problem to interaction between layers of adsorbed but mobile counterions neutralizing surfaces of similar charge density. Both when the counterion distribution is in the two-dimensional limit, and when it has a more realistic three-dimensional character, Coulomb repulsion between counterions produces an alternation of positive and negative charges at the surface. Two such apposing patterns adjust complementarily to each other, resulting in electrostatic attraction of the surfaces. The magnitude of this attraction depends solely on the surface charge density and the solution dielectric constant, while its range is defined by the size of the planar correlation hole around each ion. The attraction is stable with respect to the disruptive influence of planar thermal motion of the ions. The theory enables construction of a universal function which, after being scaled with the appropriate parameters of the system, yields the attractive electrostatic correlation pressure.


Duguid, J.G., Bloomfield, V.A., Benevides, J.M., Thomas, G.J. Jr. (1996) DNA melting studied by differential scanning calorimetry and Raman spectroscopy. Biophys. J. 71: 3350-3360.

We have used differential scanning calorimetry (DSC) and Raman difference spectroscopy to characterize the thermal melting of 160 bp fragments of calf thymus DNA at 55 mg/ml in 5 mM sodium cacodylate buffer, pH 6.36. DSC gives Tm =75.5 C, calorimetric enthalpy Hcal = 6.7 kcal/mol bp, van't Hoff enthalpy HvH = 75.6 kcal/mol bp, and entropy Scal = 21.1 cal/K mol bp. The average cooperative melting unit contains <nmelt> = 11.3 base pairs. Raman difference spectroscopy detected perturbations of marker bands for DNA secondary structure from 600 cm-1 to 1750 cm-1, during DNA melting over the range from 20 to 93 C. Increased torsional flexibility about the glycosidic bond, backbone disorder, base unstacking, and rupture of hydrogen bonds are highly correlated during DNA melting. Some new structural perturbations were detected. These include (i) base-stacking rearrangements of purine-rich bands (1300-1450 cm-1), prior to and near the end of the melting transition; (ii) shifts to lower frequencies of bands at 781 cm-1 (dC) and 1489 cm-1 (dG,dA), a direction opposite to the shifts observed upon direct metal binding; and (iii) significant decrease in intensity of the phosphodioxy (PO2-) symmetric stretch at 1092 cm-1, due to the sensitivity of the ionic phosphate group to altered hydration during melting . Two other Raman bands, at 785 cm-1 and 1014 cm-1, are recommended in lieu of the 1092 cm-1 band as standards for intensity normalization. Raman bands at 834 cm-1 (OPO) , 1240 cm-1 (dT) and 1668 cm-1 (C=O groups of dT, dG, and dC), the first sensitive to backbone geometry and the latter two to base stacking and pairing respectively, have been shown by comparison with DSC to be quantitative monitors of DNA melting. Tm's and HvH's obtained from I834, I1240 and I1668 agree well with those measured by DSC.


Last updated 5/27/98

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