The pH and Concentration Dependence of Protein-Protein Interactions, Conformation, and Viscosity in Crowded Protein Solutions

Proteins are complex macromolecules with dynamic conformations that are pH- and concentration-dependent. These attributes have marked effects on solution thermodynamics and hydrodynamics (i.e. intermolecular interactions, diffusivity, and viscosity). While significant progress has been made in dilute solution hydrodynamics and thermodynamics of proteins, there is a considerable gap in our understanding of how they are altered at high concentrations. In this talk, we attempt to bridge this gap, where we present comprehensive studies of the pH and concentration dependence of conformation and viscosity of a globular protein, Bovine Serum Albumin (BSA), and an IgG1 using small-angle neutron scattering (SANS) and microfluidic rheometry, respectively. We investigate a wide protein concentration and pH range of 2 mg/mL ≤ [Protein] ≤ 500 mg/mL and 3.0 ≤ pH ≤ 11 and find clear evidence for pH and concentration dependencies of conformation from our SANS data. The data are successfully modeled using the random-phase approximation (RPA), where we use a phenomenological model for the form factor that is able to capture contributions from both monomers and clusters in solution. Owing to the separability of the form factor into contributions from monomers and clusters, we are able to obtain structure factors that reflect monomer-monomer, monomer-cluster, and cluster-cluster correlations in solution, which allows us to gain realistic insights into packing and intermolecular interactions at high concentrations. We use these data as inputs into a modification of the model developed by Minton for protein mixtures, which can accurately capture the contributions of monomer and clustered species to model the concentration and pH dependent viscosity of BSA and the IgG1 solution.