DNA intercalation of methylene blue and quinacrine: new insights into base and sequence specificity from structural and thermodynamic studies with polynucleotides

Abstract

The binding of the known DNA intercalators methylene blue and quinacrine with four sequence specific polynucleotides, viz. poly(dG-dC).poly(dG-dC), poly(dG).poly(dC), poly(dA-dT).poly(dA-dT) and poly(dA).poly(dT), have been compared using absorbance, fluorescence, competition dialysis and thermal melting and the thermodynamic aspects of the interaction studied. In all the cases, non-cooperative binding phenomena obeying neighbor exclusion principle was observed though the affinity was remarkably higher for quinacrine and the nature of the binding was characterized to be true intercalation. The data on the salt dependence of binding derived from the plot of log Kvs. log[Na⁺] revealed a slope of around 1.0, consistent with the values predicted by the theories for the binding of monovalent cations, and contained contributions from polyelectrolytic and non-polyelectrolytic forces. The bindings were characterized by strong stabilization of the polynucleotides against thermal strand separation in both optical melting as well as differential scanning calorimetry studies. The data analyzed from the thermal melting and isothermal titration calorimetry studies were in close proximity to those obtained from absorption spectral titration data. Isothermal titration calorimetry results revealed the bindings to poly(dG-dC).poly(dG-dC), poly(dG).poly(dC) and poly(dA-dT).poly(dA-dT) to be exothermic and favoured by both negative enthalpy and large favourable positive entropy changes, while that to poly(dA).poly(dT) was endothermic and entropy driven. The heat capacity changes obtained from temperature dependence of enthalpy gave negative values to all polynucleotides. New insights on the molecular aspects of interaction of these molecules to DNA have emerged from these studies.