Mechanistic Insights into the Formation of Complex Coacervates in Crowded Environments.

Complex coacervates are widely used as model systems for studying biomolecular condensate formation, yet how intracellular crowding influences coacervation mechanisms remains unclear. Here, we investigate a model protein-polymer coacervate system in dilute and crowded environments to elucidate its formation mechanism. In dilute solutions, protein-polymer complexes gradually grow into larger assemblies as their net charge approaches neutrality. Under crowded conditions, excluded-volume effects force polymer chains into highly coiled conformations, creating a dense polymer network before the protein is introduced. Protein incorporation into this network relieves conformational tension and triggers the formation of protein-polymer networks that act as scaffolds for coacervates, bypassing the gradual charge-driven assembly seen under dilute conditions. Despite this mechanistic difference, crowders are found to be excluded from the dense phase, yielding comparable mesoscale structures under both conditions. Overall, this work suggests how condensates may adapt their assembly mechanisms within complex cellular environments.