Protein Interactions Studied by SAXS:  Effect of Ionic Strength and Protein Concentration for BSA in Aqueous Solutions

Abstract

We have studied a series of samples of bovine serum albumin (BSA) solutions with protein concentration, c, ranging from 2 to 500 mg/mL and ionic strength, I, from 0 to 2 M by small-angle X-ray scattering (SAXS). The scattering intensity distribution was compared to simulations using an oblate ellipsoid form factor with radii of 17 x 42 x 42 A, combined with either a screened Coulomb, repulsive structure factor, SSC(q), or an attractive square-well structure factor, SSW(q). At pH = 7, BSA is negatively charged. At low ionic strength, I < 0.3 M, the total interaction exhibits a decrease of the repulsive interaction when compared to the salt-free solution, as the net surface charge is screened, and the data can be fitted by assuming an ellipsoid form factor and screened Coulomb interaction. At moderate ionic strength (0.3-0.5 M), the interaction is rather weak, and a hard-sphere structure factor has been used to simulate the data with a higher volume fraction. Upon further increase of the ionic strength (I >or= 1.0 M), the overall interaction potential was dominated by an additional attractive potential, and the data could be successfully fitted by an ellipsoid form factor and a square-well potential model. The fit parameters, well depth and well width, indicate that the attractive potential caused by a high salt concentration is weak and long-ranged. Although the long-range, attractive potential dominated the protein interaction, no gelation or precipitation was observed in any of the samples. This is explained by the increase of a short-range, repulsive interaction between protein molecules by forming a hydration layer with increasing salt concentration. The competition between long-range, attractive and short-range, repulsive interactions accounted for the stability of concentrated BSA solution at high ionic strength.

Hydrogen exchange mass spectrometry reveals protein interfaces and distant dynamic coupling effects during the reversible self-association of an IgG1 monoclonal antibody

“…52 Similar trends in viscosity in response to changes in solution conditions that we describe here for mAb-C have been reported for other IgG1 mAbs, where the extent of viscosity increased in a concentration-dependent manner with increasing ionic strength 11,18,30,53,54 and solution pH, 11,55 related to elevated levels of protein RSA due to charge shielding effects. [56][57][58] The isoelectric point (pI) range of mAb-C is basic (pI»9.1-9.4), 45 and therefore, the overall surface charge of mAb-C is expected to be positive at neutral pH. As the pH was changed from 6 to 8, the tendency of mAb-C to self-associate increased, possibly due to overall decrease in electrostatic repulsive interactions.…”

Freezing lines of colloidal Yukawa spheres. I. A Rogers-Young integral equation study

“…Well-studied experimental systems which fall into this category are (sulfonate) polystyrene latex spheres in water 8,9 or in an ethanol-water mixture, 3,4 silica and polymethylacrylate (PMMA) spheres in an organic solvent (mixture), 6,10,11 and to some extent also charged globular proteins in water such as apoferritin 12,13 or bovine serum albumin (BSA). 14,15 The pair potential in Eq. (1) depends on four dimensionless parameter groups {L B /σ , Z , C s σ 3 , φ}, which can be controlled experimentally to a larger extent.…”

Study on the thermodynamics of bovine serum albumin aqueous solutions: experiments, modeling and molecular simulations.

“…However, in Zhang et al (ZHANG et al, 2007) and in Barbosa et al (BARBOSA et al, 2010), the attractive parameter (either the depth from the square well potential or the Yukawa parameter) increases when the protein concentration decreases, even though the salt concentration remains constant.…”

“…We left intentionally ε as a function of η due to the experimental evidence that ε is not constant for proteins in aqueous solutions (ZHANG et al, 2007;BARBOSA et al, 2010). As Z must approach 1 as η approaches 0, the expression for ε must follow the limit:…”

“…Previous works (BARBOSA et al, 2010;ZHANG et al, 2007) showed that the attractive parameter of proteins in solution may depend on protein concentration.…”