Proteome-scale quantification of the interactions driving condensate formation of intrinsically disordered proteins

Cellular organization in the form of biomolecular condensates is a fundamental regulatory mechanism across all forms of life. Formation of condensates relies on multivalent interactions that are often weak and transient, making them challenging to study experimentally. We have developed Condensate Partitioning by mRNA-Display (CPmD) to measure these interactions from the partition free energies of peptides and nucleic acids into reconstituted condensates. CPmD increases experimental throughput by several orders of magnitude, and we apply it to reveal the interactions driving condensate formation of intrinsically disordered proteins. We show that the partition free energies of about one hundred thousand peptides derived from the disordered proteome into a model condensate directly reflect their intrinsic propensity to form condensates. We reveal that amino acid content, linked to hydrophobicity, is the primary determinant of phase behavior. Additionally, CPmD uniquely resolves subtle sequence-dependent contributions that can encode specificity. CPmD thus provides a powerful tool to decipher how weak interactions between protein and RNA regulate biological function through condensate formation.