Thionine interaction to DNA: comparative spectroscopic studies on double stranded versus single stranded DNA

Area of Science:

• Biophysical Chemistry
• Molecular Biology
• Spectroscopy

Background:

• Thionine is a fluorescent dye with potential applications in molecular biology.
• Understanding dye-DNA interactions is crucial for developing diagnostic and therapeutic tools.

Purpose of the Study:

• To investigate the binding mechanism and affinity of thionine with double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA).
• To elucidate the role of electrostatic forces and thermodynamic parameters in thionine-DNA interactions.

Main Methods:

• Spectroscopic techniques (absorbance, fluorescence, circular dichroism)
• Equilibrium dialysis
• Isothermal titration calorimetry
• Fluorescence quenching studies

Main Results:

• Thionine exhibited strong fluorescence quenching and high binding affinity (10^5 M^-1) to dsDNA, indicative of intercalation.
• Binding to ssDNA showed weaker fluorescence quenching and lower affinity.
• Ferrocyanide quenching and polarization studies confirmed intercalative binding to dsDNA.
• Salt dependence indicated a significant role of electrostatic forces in the binding process.
• Calorimetry revealed enthalpy-entropy favorability for dsDNA binding and entropy favorability for ssDNA binding.

Conclusions:

• Thionine intercalates into dsDNA and stacks onto ssDNA, with distinct binding affinities and mechanisms.
• Electrostatic interactions contribute to the binding process.
• Thermodynamic profiles differ between dsDNA and ssDNA binding, highlighting conformational specificity.