Azure A Dye Interaction with Double-Stranded DNA Can Be Modulated by the Ionic Strength: Choosing the Binding Mode

We study the interaction of the Azure A dye with double-stranded DNA in phosphate-based buffers under three different ionic strengths, showing that the effective binding mode can be modulated by this characteristic of the surrounding buffer. For relatively high ionic strengths ([Na⁺] ≳ 10 mM), groove binding is the preferable mode, while intercalation dominates for [Na⁺] ≲ 1 mM. Single-molecule force spectroscopy assays were performed to determine these binding modes and also to extract the relevant binding parameters of the interaction in each case, resulting in a robust characterization of the complexes formed under different experimental conditions. Furthermore, gel electrophoresis assays were also performed to confirm the main conclusion about the binding mode changes. The present work advances in the characterization of DNA interactions with complex ligands, bringing new insights into the rational development of new synthetic nucleic acid dyes and drugs.