Capabilities of SEC-SAXS Method in Research of Polydisperse Systems: The Case of Apoferritin

Abstract

Here we explore capabilities of size exclusion chromatography small-angle X-ray scattering method in case of horse spleen apoferritin solution as an example of a system with high oligomerization heterogeneity. For interpretation of the experimental data, we considered models of polydisperse solution containing monomers, dimers, and trimers of apoferritin globules. Dimers were defined only by intercenter distance L, while trimers were characterized by both L and apical corner value φ. We showed that a reasonable description of the major part of the chromatogram could be reached using the model containing monomers, dimers and one type of trimers with φ about 110°–120°. In the case of two types of trimers in the model, optimal φ values were 110°–140° and 90°–110°; however, the approximation results were considered as ambiguous. Validation of the preferred model (L = 12.5 nm, φ = 120°) showed insufficient description of the experimental data for the left part of the chromatogram (elution volume <12.2 mL) indicating the presence of aggregates in these fractions. Application of the method provided the opportunity to calculate concentrations for oligomers consisting of less than four apoferritin globules, and select averaging range for different fractions properly.