A Dimer-Seeded Pathway Governs the Nucleation of ProTα-H1 Biomolecular Condensates

Abstract

Condensation between highly charged, intrinsically disordered proteins underlies physiologically essential processes such as chromatin regulation and nucleolus self-assembly, yet the mechanism by which these condensates first nucleate from the dilute phase has remained essentially inaccessible to both experiment and atomistic simulation. Here we use residue-level coarse-grained simulations of complex coacervates formed by Prothymosin (ProT) with histone H1, protamine, poly-lysine and poly-arginine, and show that they recapitulate the experimental salt-dependent phase behaviour, binding affinities, chain reconfiguration, protein diffusion and viscosity of these systems. Combining dilute-phase binding free energies with brute-force nucleation simulations, we reveal that these condensates assemble through a dimer-seeded nucleation mechanism: oppositely charged, polyanionic and polycationic partners first pair into a charge-compensated dimer--the most stable minimal complex and the dominant pre-nucleation species--from which clusters subsequently grow into large-scale condensates. The free energy cost of forming these clusters rises with salt and becomes density-controlled for larger assemblies, placing nucleation under the joint control of electrostatic screening and protein supersaturation. This same electrostatic screening shapes mature mesoscopic condensates: contacts form and break within picoseconds regardless of the phase, while chain reconfiguration and diffusion slow down markedly upon condensation, so that a condensate is set apart not by the strength of its contacts but by their collective network connectivity.