We have set a goal to investigate the biophysical and structural basis of the interaction between interferons and their receptors using purified proteins. Using site- directed mutagenesis and biophysical measurements we have mapped the binding sites of IFNoo 2 and IFNoooo on ifnar2, as well as the binding site of ifnar2 on IFNoo 2. Comparing the binding epitope of IFNoo 2 and IFNoooo on ifnar2 revealed that the two IFNs have distinct binding centers, which might be responsible for the differences in transmembrane activation observed between the two interferons. The structural and thermodynamic basis of this interactions was further investigated using double- mutant cycles. These data were incorporated into a newly developed docking algorithm, which enables us to build a structural model of the complex.
Further studies are currently underway to improve the quality of our model.
This method will be applied to address another goal, namely the understanding of the mode of interaction of different type I IFNs with their common receptors. This will be done by mapping the mode of interaction of the ternary complex - ifnar1, ifnar2 and interferon, to elucidate structural, kinetic and thermodynamic differences in binding of different interferon subtypes. To complement these biophysical studies with high resolution structural data we are searching for crystallization conditions for the IFNoo 2- ifnar2 complex. We aim to correlate structural, thermodynamic and kinetic data of binding with the interferon mediated cellular response of the different type I interferon subtypes. Our long term goal is to be able to use this gained understanding for the design of interferons, or interferon- like molecules with improved specific activity in curing disease.