Sorption, Intercalation and Cooperativety: the Modes of Interaction of Actinomycin to DNA

Abstract

The interaction of Actinomycin-D to DNA has been long investigated given it inhibits the synthesis of ribonucleic acid, inhibits the growth of cancer cells and induces apoptosis. So far, thermodynamic and structural studies have demonstrated that Actinomycin-D intercalates to DNA double helix preferentially to G-C pairs. There is also evidence that binding affinity is modulated by nearest base pairs flanking the intercalation site. However, the mechanism of Actinomycin-D interaction to DNA, and thus its energetic, is still ill understood. While some studies show evidence that ActD intercalation to natural DNA occurs via a mechanism consistent with a model of one independent and equivalent sites, other studies show evidence of the existence of two classes of independent binding sites; other yet show that the binding at low saturation is cooperative. In this work we measured the binding of Actinomycin-D to calf thymus DNA by optical titration and dialysis equilibrium under different solution conditions. Thus, we have found the conditions where the different kinds of binding reported in the literature can be reproduced. Through the analysis of the data correlating experimental design and solutions conditions, we were able to characterize the complexity of ActD interactions with DNA. In this work we show experimental evidences that intercalation of ActD at low drug/DNA ratio is cooperative; that the strong binding site is a consequence of cooperative binding; and that ActD not only intercalates to the DNA double helix but it also binds to the helix surface with a affinity which is in the same order of that measured upon intercalation.