Excess counterion accumulation around branched nucleic acids

Many nucleic acids of biological importance possess elements of tertiary structure in which the regional phosphate charge density dramatically exceeds that of linear duplex DNA, as in the inter-helix junctions found in tRNAs, ribosomal RNAs, and Holliday intermediates in general recombination. However, despite a long-standing awareness that such structures have special counterion requirements for stability few studies have focused on their level of counterion association. In order to gauge the influence of high regional phosphate charge density on the extent of counterion association, we have defined the degree of "excess counterion association" for a four-branch DNA junction as the number of additional counterions (over the relevant linear DNA value) that are associated with the junction in a Donnan equilibrium dialysis experiment. Grand canonical Monte Carlo computations were used to determine the Donnan distribution (preferential interaction) coefficients, employing a "primitive model" description of the nucleic acid and the 1:1 electrolyte. We have determined that at least 24 excess counterions are associated with the junction in the long branch limit. The subsequent release of a portion of these additional counterions during the process of ligand binding is therefore likely to provide a strong directional influence on the binding of proteins and cationic ligands, with preferred binding near or on the junction vertex or near other elements of tertiary structure (e.g. pseudo-knots or triplexes) even if the ligands do not directly recognize the structural elements themselves. Moreover, excess counterion association is expected to play a significant role in determining the relative stabilities of alternative tertiary structures.