Identification of a PU.1–IRF4 protein interaction surface predicted by chemical exchange line broadening

Relaxation values reflecting residue-specific line broadening revealed amino acids in the DNA-binding domain of PU.1 on a surface potentially involved in protein–protein interactions. Mutation of these amino acids did not cause protein unfolding but destabilized PU.1–DNA binding. Addition of IFN response factor 4 to form the ternary complex recovered binding stability. Fluorescence quenching experiments proved that this surface of PU.1 interacts with IFN response factor 4 during binding. Our results provide evidence that residues that display increased conformational exchange can be used to predict areas of protein–protein interactions.