Interactions in protein solutions close to liquid-liquid phase separation: Ethanol reduces attractions via changes of the dielectric solution properties

TL;DR

This study investigates how ethanol and NaCl influence protein phase separation and interactions in lysozyme solutions, revealing that ethanol reduces attractions by altering dielectric properties, with results supported by multiple experimental methods.

Contribution

The paper provides a quantitative analysis of how ethanol and salt modify protein interactions and phase behavior through dielectric effects, using the extended law of corresponding states and DLVO theory.

Findings

Ethanol lowers the cloud-point temperature, indicating reduced protein attraction.

NaCl raises the cloud-point temperature, indicating increased attraction.

Calculated $B_2$ values agree with experimental scattering data and theoretical models.

Abstract

Ethanol is a common protein crystallization agent, precipitant, and denaturant, but also alters the dielectric properties of solutions. While ethanol-induced unfolding is largely ascribed to its hydrophobic parts, its effect on protein phase separation and inter-protein interactions remains poorly understood. Here, the effects of ethanol and NaCl on the phase behavior and interactions of protein solutions are studied in terms of the metastable liquid-liquid phase separation (LLPS) and the second virial coefficient $B_2$ using lysozyme solutions. Determination of the phase diagrams shows that the cloud-point temperatures are reduced and raised by the addition of ethanol and salt, respectively. The observed trends can be explained using the extended law of corresponding states as changes of $B_2$. The results for $B_2$ agree quantitatively with those of static light scattering and small-angle X-ray scattering experiments. Furthermore, $B_2$ values calculated based on inter-protein interactions described by the Derjaguin--Landau--Verwey--Overbeek (DLVO) potential and considering the dielectric solution properties and electrostatic screening due to the ethanol and salt content quantitatively agree with the experimentally observed $B_2$ values.