Liquid phase separation controlled by pH

TL;DR

This paper introduces a minimal thermodynamic model to understand how pH influences liquid phase separation in macromolecular mixtures, highlighting the role of charge states and predicting reentrant phase behavior.

Contribution

The study develops a thermodynamic framework linking pH-dependent protonation reactions to phase separation, providing insights into protein behavior near the isoelectric point.

Findings

Phase diagrams vary with pH and charge interactions.

Reentrant phase separation behavior predicted as pH changes.

Broader phase separation region at the isoelectric point.

Abstract

We present a minimal model to study liquid phase separation in a fixed pH ensemble. The model describes a mixture composed of macromolecules that exist in three different charge states and have a tendency to phase separate. We introduce the pH dependence of phase separation by means of a set of reactions describing the protonation and deprotonation of macromolecules, as well as the self-ionisation of water. We use conservation laws to identify the conjugate thermodynamic variables at chemical equilibrium. Using this thermodynamic conjugate variables we perform a Legendre transform which defines the corresponding free energy at fixed pH. We first study the possible phase diagram topologies at the isoelectric point of the macromolecules. We then show how the phase behavior depends on pH by moving away from the isoelectric point. We find that phase diagrams as a function of pH strongly depend on whether oppositely charged macromolecules or neutral macromolecules have a stronger tendency to phase separate. We predict the existence of reentrant behavior as a function of pH. In addition, our model also predicts that the region of phase separation is typically broader at the isoelectric point. This model could account for both, the protein separation observed in yeast cells for pH values close to the isoelectric point of many cytosolic proteins and also for the in vitro experiments of single proteins exhibiting phase separation as a function of pH.