Department of Biochemistry, Sydney University
Recent developments in analytical ultracentrifugation, both in instrumentation and methods of analysis, have revived this method as a means of characterisation of macromolecules. While sequencing and mass spectrometry provide ultimate precision in determination of the molecular weights of biological polymers, analytical ultracentrifugation is specially suited to determining the molecular weight and molecular weight distributions in solution. In particular, analytical ultracentrifugation in sedimentation equilibrium experiments can provide information on the stoichiometry and thermodynamics of reversible interactions involving macromolecules: binding of small ligands, self-association, and heterogeneous associations such as DNA-protein interactions.
Sedimentation equilibrium experiments also provide the only reliable means of studying association reactions at equilibrium, without perturbation, and provide a means for checking whether equilibrium has been attained and whether the solution is free of non-participating contaminants such as the products of irreversible aggregation or proteolysis.
Sedimentation velocity and diffusion measurements in the ultracentrifuge provide additional information on the approximate sizes and shapes of solute molecules, and can provide further information on the thermodynamics and kinetics of interaction behaviour.