Abstract
The reversible formation of protein-protein interactions plays a crucial role in many biological processes. In order to carry out a thorough quantitative characterization of these interactions, it is essential to establish the oligomerization state of the individual components first. The sedimentation equilibrium method is ideally suited to perform these studies because it allows a reliable, accurate, and absolute value of the solution molecular weight of a macromolecule to be obtained. This technique is independent of the shape of the macromolecule under investigation and allows the determination of equilibrium constants for a monomer-multimer self-associating system.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Minton, A. P. (1990) Quantitative characterization of reversible molecular associations via analytical centrifugation. Anal. Biochem. 190, 1–6.
McRorie, D. K. and Voelker, P. (1993) Self-associating systems in the analytical ultracentrifuge, Beckman, Fullerton, CA.
Rivas, G. and Minton, A. P. (1993) New developments in the study of biomolecular associations via sedimentation equilibrium. Trends Biochem. Sci. 18, 284–287.
Hansen, J. C., Lebowitz, J., and Demeler, B. (1994) Analytical ultracentrifugation of complex macromolecular systems. Biochemistry 33, 13,155–13,163.
Schuster, T. M. and Toedt, J. M. (1996) New revolutions in the evolution of analytical ultracentrifugation. Curr. Opin. Struct. Biol. 6, 650–658.
Cole, J. L. and Hansen, J. C. (1999) Ananlytical ultracentrifugation as a contemporary biomolecular research tool. J. Biomol. Techn. 10, 163–176 (URL: http://www.beckman.com/resourcecenter/literature/BioLit/BioLitList.asp).
Laue, T. M. and Stafford, W. F., 3rd (1999) Modern applications of analytical ultracentrifugation. Annu. Rev. Biophys. Biomol. Struct. 28, 75–100.
Liu, J. and Shire, S. J. (1999) Analytical ultracentrifugation in the pharmaceutical industry. J. Pharm. Sci. 88, 1237–1241.
Cantor, C. R. and Schimmel, P. R. (1980) Biophysical Chemistry, Part II, Freeman, San Francisco.
Laue, T. (1996) Choosing which optical system of the Optima XL-I analytical centrifuge to use, Beckman-Coulter Technical Application Information Bulletin A-1821-A (URL: http://www.beckman.com/resourcecenter/literature/BioLit/BioLitList.asp).
Johnson, M. L., Correia, J. J., Yphantis, D. A., and Halvorson, H. R. (1981) Analysis of data from the analytical ultracentrifuge by nonlinear least-squares techniques. Biophys. J. 36, 575–588.
Johnson, M. L. and Faunt, L. M. (1992) Parameter estimation by least-squares methods. Methods Enzymol. 210, 1–37.
Sedgwick, S. G., Taylor, I. A., Adam, A. C., et al. (1998) Structural and functional architecture of the yeast cell-cycle transcription factor swi6. J. Mol. Biol. 281, 763–775.
Binns, D. D., Barylko, B., Grichine, N., et al. (1999) Correlation between self-association modes and GTPase activation of dynamin. J. Protein Chem. 18, 277–290.
Binns, D. D., Helms, M. K., Barylko, B., et al. (2000) The mechanism of GTP hydrolysis by dynamin II: a transient kinetic study. Biochemistry 39, 7188–7196.
Silkowski, H., Davis, S. J., Barclay, A. N., Rowe, A. J., Harding, S. E., and Byron, O. (1997) Characterisation of the low affinity interaction between rat cell adhesion molecules CD2 and CD48 by analytical ultracentrifugation. Eur. Biophys. J. 25, 455–462.
Rivas, G., Stafford, W., and Minton, A. P. (1999) Characterization of heterologous protein-protein interactions using analytical ultracentrifugation. Methods 19, 194–212.
Philo, J. S. (2000) Sedimentation equilibrium analysis of mixed associations using numerical constraints to impose mass or signal conservation. Methods Enzymol. 321, 100–120.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2004 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Taylor, I.A., Eccleston, J.F., Rittinger, K. (2004). Sedimentation Equilibrium Studies. In: Fu, H. (eds) Protein-Protein Interactions. Methods in Molecular Biology, vol 261. Humana Press. https://doi.org/10.1385/1-59259-762-9:119
Download citation
DOI: https://doi.org/10.1385/1-59259-762-9:119
Publisher Name: Humana Press
Print ISBN: 978-1-58829-120-2
Online ISBN: 978-1-59259-762-8
eBook Packages: Springer Protocols