Binding affinities on all scales from fragment screening to intrinsically disordered proteins with isothermal spectral shift technology

The precise characterisation of biomolecular interactions, their binding constants and stoichiometries, lie at the heart of understanding events in cells and disease. Furthermore, the discovery and development of new drugs often requires the screening of chemical libraries containing hundreds if not many thousands of chemical compounds. After a small number of hit compounds are discovered, the optimisation towards a promising lead compound is greatly aided by a precise knowledge of their binding to their biological targets. Among the many different screening technologies, biophysical techniques that precisely measure the strength of the binding (binding constant Kd) and the composition of the complex (stoichiometry) are often considered the gold standard. However, many therapeutic targets are challenging and some are still entirely within the reach of current biophysical techniques. These include many membrane proteins that cannot be produced at the required amount or concentration. The Dianthus by NanoTemper Technologies, based on a novel spectral shift technology, represents a huge leap in sensitivity which enables the precise measurement of binding constants in very small volumes (less than five microliter) and at very low (nanomolar) concentrations. Hence, analyses of the most challenging protein targets, such as membrane proteins that often require detergents and intrinsically disordered proteins that tend to precipitate even at moderate concentrations, become feasible. Furthermore, combining this technology with high-throughput methods allows the measurements of 384 wells in parallel in less than one hour, which allows the screening of entire chemical libraries with thousands of compounds. Importantly, the Dianthus system will be the centre piece of the biophysical screening platform at the Biophysical Sciences Institute at Durham University which includes a wide range of complementary instrumentation. Taken together with the technical and scientific...