Method of measuring oligonucleotide-metal affinities: Interactions of the thrombin binding aptamer with K+ and Sr2+

摘要

We report a new, mass spectrometry-based method for measuring affinity constants for specific metal ion binding to DNA, particularly for quadruplex DNA. This method, which is applicable to other systems, utilizes the gas-phase signal fractions, as determined by mass spectrometry, from the bound and unbound species as input into a mathematical model that determines various parameters, one of which is the binding affinity constant. The system used to develop and test the model was the thrombin-binding aptamer, an appropriate quadruplex structure that binds both K+ and Sr2+ cations. Using this method, we measured the binding constants of potassium and strontium cations with the quadruplex structure to be 5000 and 240 nM, respectively. We then applied the method to measure the change in enthalpy of the binding of strontium cations to the thrombin binding aptamer. The Delta H for this interaction is -71 kJ/mol (-17 kcal/mol). The binding constant measurements are consistent with earlier measurements on the same system, and the measured change in enthalpy is in excellent agreement with previous work.