Allosteric conversion of Z DNA to an intercalated right-handed conformation by daunomycin

Absorbance and fluorescence methods were used to measure the binding of the anticancer drug daunomycin to poly (dGdC) under ionic conditions that initially favor the left-handed Z conformation of the polymer. Drug binding was cooperative under these conditions and may be fully accounted for by an allosteric model in which the drug binds preferentially (but not exclusively) to the right-handed B conformation and shifts the polymer from the Z to an intercalated right-handed conformation. Quantitative analysis of binding isotherms in terms of the allosteric model allowed for estimation of the equilibrium constants for the conversion of a base pair at a B-Z interface from the Z to the B conformation and for the formation of a base pair in the B conformation within a stretch of helix in the Z conformation. The free energy of the Z to B conversion of a base pair was calculated from this data and ranges from +0.03 to +0.3 kcal/mol over the NaCl range of 2.4-3.5 M. The free energy for the formation of a B-Z junction was nearly constant at +4.0 kcal/mol over the same range of NaCl concentrations. The salt dependence of the free energy of the Z to B transition indicates preferential Na+ binding to the Z form and that there is a net release of Na+ upon conversion of a base pair from the Z to the B conformation. The energetically unfavorable Z to B transition was found by this analysis to be driven by coupling to the energetically favorable interaction of daunomycin with B form DNA. In 3.5 M NaCl, for example, the free energy change for the overall reaction (Z DNA base pairs) + (daunomycin) in equilibrium with (right-handed complex) is -7.0 kcal/mol, nearly all of which is contributed by the binding of drug to B DNA. Analysis using the allosteric model also shows that the number of base pairs converted from the Z to the B conformation per bound drug molecule is salt dependent and provides evidence that drug molecules partition into regions of the polymer in the right-handed conformation.