Analytical ultracentrifugation and size- exclusion chromatography coupled with light scattering for the characterization of membrane proteins in solution

Abstract

Analytical ultracentrifugation (AUC) sedimentation velocity (SV) with absorbance and interference detection and size-exclusion chromatography coupled with static and dynamic light scattering, absorbance, and refractive index detections (SEC/MALS) are two techniques that combine separation and analysis, in an absolute manner, of the mass and size of macromolecules in solution. We present here how they can be applied to the study of membrane proteins. We describe briefly the principles of the species separation, what the detection systems used measures, some theoretical background, the steps for data analysis with the example of the outer membrane protein FhuA, and emphasize the complementarity between these two techniques.