AZURE A DYE INTERACTION WITH DNA: A SINGLE MOLECULE FORCE SPECTROSCOPY STUDY

This work presents, for the first time, the investigation of the interaction between double-stranded DNA and the Azure A dye at the single molecule level, using the optical tweezers technique. Azure A is a phenothiazine dye of interest due to its diverse applications in areas such as labeling and photosensitization, where the understanding of its interactions with biomolecules is fundamental. The interaction between DNA and Azure A was investigated here under three different ionic strengths: [Na+] = 150, 10, and 1 mM. Using optical tweezers to perform single molecule force spectroscopy, we were able to identify the direct dependence of the binding modes with the ionic strength of the surrounding buffer. More significantly, we were able to quantify key physicochemical (binding) parameters such as the equilibrium binding constant and the exclusion number (binding site size), derived from measurable changes on the mechanical properties of the DNA-dye complexes (contour persistence lengths) upon dye binding. These parameters, inherent to the mechanics of polymers and sensitive to structural changes, provided detailed results on the nature of the binding and how the presence of Azure A modulates the elasticity and conformation of DNA in vitro. The data obtained in this study significantly deepens our understanding of the complex molecular interactions between dyes and nucleic acids and demonstrate that we can use Azure A as a DNA binding agent for various applications.
This work was supported by Conselho Nac. Des. Cient. Tecnológico (CNPq # 305185/2022-5) and by Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG # APQ-04822-23).