Enhanced protein adsorption upon bulk phase separation

Madeleine R Fries,Fajun Zhang,Robert M J Jacobs,Alexander Hinderhofer,Maximilian W A Skoda,Christoph Kertzscher,Daniel Stopper,Matthias Blum,Frank Schreiber,Roland Roth

doi:10.1038/s41598-020-66562-0

Abstract

In all areas related to protein adsorption, from medicine to biotechnology to heterogeneous nucleation, the question about its dominant forces and control arises. In this study, we used ellipsometry and quartz-crystal microbalance with dissipation (QCM-D), as well as density-functional theory (DFT) to obtain insight into the mechanism behind a wetting transition of a protein solution. We established that using multivalent ions in a net negatively charged globular protein solution (BSA) can either cause simple adsorption on a negatively charged interface, or a (diverging) wetting layer when approaching liquid-liquid phase separation (LLPS) by changing protein concentration (cp) or temperature (T). We observed that the water to protein ratio in the wetting layer is substantially larger compared to simple adsorption. In the corresponding theoretical model, we treated the proteins as limited-valence (patchy) particles and identified a wetting transition for this complex system. This wetting is driven by a bulk instability introduced by metastable LLPS exposed to an ion-activated attractive substrate.

Highlights

In all areas related to protein adsorption, from medicine to biotechnology to heterogeneous nucleation, the question about its dominant forces and control arises
We study the adsorption of an aqueous bovine serum albumin (BSA) solution in the presence of YCl3 at a silicon dioxide (SiO2) interface[44] as a function of cs experimentally by means of ellipsometry and quartz-crystal microbalance with dissipation (QCM-D) and theoretically within the framework of classical density functional theory (DFT)
Our experimental and theoretical results suggest that the enhanced protein adsorption upon approaching liquid-liquid phase separation (LLPS) features the onset of a ‘wetting’ transition caused by dominatingly attractive protein-protein and protein-substrate interactions, both of which are mediated by the multivalent ions

Summary

Introduction

In all areas related to protein adsorption, from medicine to biotechnology to heterogeneous nucleation, the question about its dominant forces and control arises. The behaviour of proteins in the presence of multivalent salt can be successfully modelled as patchy colloids, where the patches are activated by cations[20]: In addition to a hard-sphere-like core repulsion, a patch-patch interaction is mediated by ions which can activate the sites by chemically binding to the protein surface; a bond between two distinct proteins is only possible if an activated patch meets a deactivated one (see Methods for further details). The resulting phase diagrams, which can be obtained from Wertheim’s perturbation theory for associating particles[40,41,42], are in excellent qualitative agreement with the experiments considering the coarse-grained nature of the model This includes RC in terms of protein clusters and a closed-loop LLPS region. This is a prominent feature of patchy fluids[43], and cannot be understood with fluids interacting via isotropic forces where liquid densities often reach volume fractions of 40% or beyond[10]

Methods

Results

Conclusion