Characterisation of Macromolecular and Ligand Interactions using SPR and ITC

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We propose to develop, on the science campus of Edinburgh University, a state-of-the-art research facility for biophysical characterisation of the binding events involved in forming biomolecular complexes. Specifically, we request equipment to measure surface plasmon resonance (SPR) and isothermal titatration calorimetry ITC data. The proposal is driven by numerous projects emanating from the University¿s very large community of biomolecular scientists spanning three 5-A rated academic departments that will benefit from the requested equipment. The seven applicants are a subset of this community who have a particularly pressing need of, and therefore a compelling case for, access to in-house SPR and ITC. The proposal involves collaboration with Biacore since we have requirements for methods development as well as routine work. The proposed work includes studies of the thermodynamics of ligand-binding and assay design aimed at lead-discovery, protein-protein interactions in forming complexes important for immunity and infection, protein- and DNA-metal interactions pertinent to the pharmaceutical industry, protein-DNA interactions involved in epigenetics, and interactions of proteins implicated in neurodegenerative diseases. These projects reflect broad strengths at Edinburgh in transitional biology, structural biology, molecular cell biology and (increasingly) in systems biology. From a strategic viewpoint, the generation of quantitative data for intermolecular interactions will foster collaborations between biologists working in key areas, and chemists, biochemists and biophysicists who need quantitative data to infer mechanisms and design inhibitors with potential commercial value. The technical and developmental aspects of the proposal exploit the sensitivity of the new Biacore T-100 SPR instrument to allow determination of the enthalpy and entropy of the transition state and also of the final protein-ligand complex. This facilitates a novel SPR-based approach to studying interactions. Critically, the data generated can be compared with parameters derived from ITC. Biacore will provide financial and technical support for these experiments. All the systems chosen for these studies have also been well characterised structurally in our laboratories which will enable a very detailed interpretation of the thermodynamic data. The external sponsors and the university provide over 33 per cent of the costs of this project. The requested instrumentation will allow us to build on the outstanding facilities already established within the Edinburgh Protein Interaction Centre (launched in 2001) and hence to maintain our internationally competitive edge.