Isothermal Titration Calorimetry for Studying Macromolecular Interactions

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The unifying theme of our NSERC DG-funded research programs is to establish the role of biomolecular interactions (interactions of proteins with nucleic acids, small molecule interactions with proteins and nucleic acids, etc.,) in cellular processes in organisms such as viruses, bacteria, yeast, and humans. The pivotal challenge we face in advancing our research and training activities is the lack of critical infrastructure we require to study biomolecular interactions. Without this, we cannot perform interdisciplinary research to train students and researchers at the UofL. Recent advances in structural biology methods have expanded our understanding of how biomolecules communicate with each other and function. However, to study the high-resolution structures of biomolecular complexes, information on affinity and stoichiometry is essential. Therefore, we propose to install isothermal titration calorimetry (ITC). This widely used and reliable instrument aids a direct measure of the heat generated or absorbed as a result of biomolecular interaction. ITC allows the direct, rapid, and accurate determination of binding affinities and it is possible to study interactions of a wide range of unaltered biomolecules in their native state. In a single experiment, ITC accurately determines a wide range of binding constants, reaction stoichiometry, as well as thermodynamic parameters such as enthalpy and entropy. In addition to binding affinities, ITC can help elucidate the mechanisms underpinning molecular interactions. The UofL does not have an ITC instrument; hence, there is an urgent need for this critical infrastructure. In conjunction with the existing infrastructure at the UofL, the installation of ITC will accelerate HQP training in our laboratories, and in our collaborator's laboratories, and significantly advance research and training activities.