Multistep Desolvation as a Fundamental Principle Governing Peptide Self-Assembly Through Liquid-Liquid Phase Separation

Biomolecular self-assembly based on peptides and proteins is a general phenomenon encountered in natural and synthetic systems.Liquid–liquid phase separation(LLPS)is intimately involved in biomolecular self-assembly,yet the key factors at a molecular scale activating or modulating such a process remain largely elusive.Herein,we discovered in our experiments that multistep desolvation is fundamental to the formation and evolution of peptide-rich droplets:The first step was partial desolvation of peptides to form peptide clusters,and the second step was selective desolvation of hydrophobic groups within clusters to trigger LLPS and the formation of peptiderich droplets,followed by complete desolvation of droplets,initiating the nucleation of peptide selfassembly.Manipulation of the degree of desolvation at different stages was an effective strategy to control the self-assembly pathways and polymorphisms.This study sheds light on the molecular origin of LLPS-mediated self-assembly distinct from classical one-step self-assembly and paves the way for the precise control of supramolecular self-assembly.