[Untitled]

Hotspots are defined as the minimal functional domains involved in protein:protein interactions and sufficient to induce a biological response. Here we describe the use of complex and high diversity phage display libraries to isolate peptides (called Hotspot Ligands or HSPLs) which sub-divide the ligand binding domain of the tumor necrosis factor receptor 2 (TNFR2; p75) into multiple hotspots. We have shown that these libraries could generate HSPLs which not only subdivide hotspots on protein and non-protein targets but act as agonists or antagonists. Using this approach, we generated peptides which were specific for human TNFR2, could be competed by the natural ligands, TNFα and TNFβ and induced an unexpected biological response in a TNFR2-specific manner. To our knowledge, this is the first report describing the dissection of the TNFR2 into biologically active hotspots with the concomitant identification of a novel and unexpected biological activity.