Quantifying the Thermodynamic Profiles of the Interaction between Small Molecules and Nucleic Acids by Single Molecule Techniques

Thermodynamic profiles of the interaction of small molecules with nucleic acid are essential to rational drug design. Accurately predicting binding affinity constant is highly required to determine the binding energetics of the driving forces in drug-DNA interactions. Superior to bulk measurements, single-molecule measurements using optical tweezers provide detailed information on the mechanical and elastic properties of double-strand DNA molecules in the presence of binding ligands. Recently, we have applied optical tweezers to investigate ligand-DNA intercalative binding study [ChemPhysChem, 2009, 10(16), 2791; Biochemical and Biophysical Research Communications, 2010]. The proposed schemes accurately predict the site exclusion number (n), binding affinity constant (KA), and change of binding free energy. On the other hand, we have designed the temperature control system that controls the microenvironment temperature ranging from 20℃ to 35℃. In addition, we can further apply the proposed single-molecule approach, together with Gibbs function (ΔG=ΔH-TΔS), to determine the thermodynamic profiles including free energy change (ΔG), enthalpy change (ΔH), and entropy change (ΔS), where G, H and S represent free energy, enthalpy and entropy, respectively. The project will present our single-molecule approach to determine the complete thermodynamic profiles for DNA binding drugs for the first time. This approach is particularly important in rational drug design, compound identification and optimization. This project will primarily focus on four specific aims, and we will outline them briefly: Aim 1: Construct single-molecule manipulation technique and establish approach to predict the micro-environmental temperature inside the microfludic system; Aim 2: Using single-molecule manipulation technique, together with fluorescence technique to determine the binding mode of the DNA binding drugs; Aim 3: Validate the DNA biomechanics due to the presence of DNA binding drugs, and Aim 4: Complete the DNA thermodynamic profiles of DNA binding drugs; this allows us to validate the second law of thermodynamics for small system.