As part of our comprehensive study of the double and triple helicies of the poly dA- poly dT system over a wide range of salt concentration, we have found that the melting temperature of the B-DNA like poly (dA + dT) double helix is not simply linear in the logarithm of either the concentration or the activity of NaCl. The behavior is more complex and is well represented by a cubic polynomial over the range of 0.01 M to 0.9 M NaCl. Differential scanning calorimetry results show that the melting enthalpy varies with salt concentration. The dependencies of the melting temperature and enthalpy upon salt concentration can be appropriately combined to yield the quantity 2 Delta Gamma. This quantity has often been interpreted to represent the net number of sodium ions released upon melting of the double helix and is generally believed to be a constant value characteristic of the stoichiometry of the helix transformation reaction. We have found that 2 Delta Gamma for poly (dA + dT) is not constant, but varies more than two-fold over the range of salt concentration examined. Combination of our values of Delta Gamma with the results of dialysis equilibrium studies of native DNA gives wholly experimental values of Gamma, the salt distribution parameter that describes the Donnan membrane equilibrium of the melted, coiling single strands of poly dA and poly dT. At any given salt concentration, solution of the non-linear Poisson - Boltzmann equation for an impenetrable cylinder of assumed charge separation and radius gives the electrostatic potential as a function of distance from the axis of the rod. Integration of this potential over the annulus from the outer boundary of the model cylinder to the mathematical infinite radial distance enables calculation of Gamma, the salt distribution parameter for the denatured single strands, as a function of ionic strength. Satisfactory agreement between the experimental values of Gamma for the single strands and the calculated curves of Gamma as a function of salt concentration has been obtained using a model of a completely dissociated stiff rod with a 4.7 angstrom radius and a charge separation projected upon the axis of the cylinder of 4.3 angstroms. This radius is about one half that of native B-form DNA, which is reasonable, while the charge separation is found to be somewhat larger than the value of 3.4 angstroms characteristic of completely stacked bases in helical DNA that has often been given as an estimate for the melted single strands. This analysis based upon three- component thermodynamics and the classical electrostatic Poisson - Boltzmann equation provides insight into the conformation and dimensions of the coiled forms in solution and gives a satisfactory account of the observed complex dependence of the double-helix melting temperature upon salt concentration.
| Ross, Philip D; Howard, Frank B (2003) The thermodynamic contribution of the 5-methyl group of thymine in the two- and three-stranded complexes formed by poly(dU) and poly(dT) with poly(dA). Biopolymers 68:210-22 |
