Expanding the molecular language of protein liquid–liquid phase separation

Understanding the relationship between a polypeptide sequence and its phase separation has important implications for analysing cellular function, treating disease and designing novel biomaterials. Several sequence features have been identified as drivers for protein liquid–liquid phase separation (LLPS), schematized as a ‘molecular grammar’ for LLPS. Here we further probe how sequence modulates phase separation and the material properties of the resulting condensates, targeting sequence features previously overlooked in the literature. We generate sequence variants of a repeat polypeptide with either no charged residues, high net charge, no glycine residues or devoid of aromatic or arginine residues. All but one of 12 variants exhibited LLPS, albeit to different extents, despite substantial differences in composition. Furthermore, we find that all the condensates formed behaved like viscous fluids, despite large differences in their viscosities. Our results support the model of multiple interactions between diverse residue pairs—not just a handful of residues—working in tandem to drive the phase separation and dynamics of condensates.