Intercalation of ethidium ion into DNA and RNA oligonucleotides.

Abstract

AbstractThe thermodynamics of ethidium ion binding to the double strands formed by the ribooligonucleotides rCA5G + rCU5G and the analogous deoxyribo‐oligonucleotides dCA5G + dCT5G were determined by monitoring the absorbance versus temperature at 260 and 283 nm at several concentrations of oligonucleotides and ethidium bromide. A maximum of three ethidium ions bind to the oligonucleotides, which is consistent with intercalation and nearest‐neighbor exclusion. For the ribo‐oligonucleotide the binding mechanism is complex. Either two sites (assumed to be the intercalation sites at the two ends of the oligonucleotide) bind more strongly by a factor of 140 than the third site, or all sites are identical, but there is strong anticooperativity on binding (cooperativity parameter, 0.1). In sharp contrast, the binding to the same sequence (with thymine substituted for uracil) in the deoxyribo‐oligonucleotide showed all sites equivalent and no cooperativity. For the ribo‐oligonucleotides the enthalpy for ethidium binding is −14 kcal/mol. The equilibrium constants at 25°C depend on the model; either K = 6 × 105M−1 for the two strong sites (4 × 103M−1 for the weak site) or K = 2.5 × 105M−1 for the intrinsic constant of the anticooperative model. For the equivalent deoxyribo‐oligonucleotide the enthalpy of binding is ‐9 kcal/mol and the equilibrium constant at 25°C is a factor of 10 smaller (K = 2.5 × 104M−1).