Phase Separation of Polypeptide Solutions: Interplay between Electrostatics and Non-electrostatic Specific Interactions

The liquid-liquid phase separation of biomacromolecules, including proteins and RNAs, plays a crucial role in the formation of many biocondensates that govern various biological functions, such as ribosome biogenesis and cellular stress response. The understanding of the underlying driving forces, especially the roles of electrostatics and weak non-electrostatic specific interactions, remains elusive. Here we develop a simple mean- field theory to systematically examine the phase behaviors of polypeptide solutions with both electrostatics and non-electrostatic specific interactions such as hydrogen bonding and pi-pi stacking. Our theory treats the electrostatic correlation via the generalized Debye-Hückel theory and the first-order thermodynamic perturbation theory for chain connectivity. The specific interaction is accounted for by the sticker-spacer model. We find that in salt-free solutions, specific interactions alone can drive phase separation when the charge fraction is low. As the charge fraction increases to moderate levels, strong electrostatic correlation is necessary to induce phase separation, with its effect initially being suppressed and then promoted with increasing the charge fraction. For salty solutions without specific interactions, phase separation occurs only when electrostatic correlation is sufficiently strong, and the miscibility gap expands with increasing the charge fraction. Introducing specific interactions lowers the electrostatic strength required to induce phase separation, while increasing the charge fraction shrinks the phase separation window. Finally, increasing the salt concentration screens the electrostatic repulsion between charged amino acids and thus leads to phase separation in solutions that are otherwise homogeneous. Our theoretical calculations are in reasonable agreement with both the experimental observations as well as our molecular dynamics simulations. We rationalize our findings via the interplay of counterion entropy, excluded volume interaction, electrostatic correlation, and specific interaction.