Protein IDR linkers regulate overall protein conformation and phase separation

The phase separation of proteins is considered to be promoted by driver regions. However, emerging evidence indicates that disordered linkers connecting driver regions in multidomain proteins may regulate phase separation. In this study, we use a protein-DNA system to investigate how disordered linkers affect protein phase separation behavior. We report that the residue-residue interactions within disordered linkers modulate the overall conformational ensemble of the protein, thereby influencing the threshold concentration for phase separation and physical properties of the resulting condensates. We demonstrate that disordered linkers not only serve a connecting role but also play a previously underexplored regulatory role in protein phase separation through their sequence diversity. This addresses a gap in the understanding of linker-mediated regulation of phase separation and advances the "stickers-and-spacers" theory. Furthermore, we show that a molecular grammar governs condensate properties, providing valuable insights into the relationship between disordered linker sequences and condensate functions.