# Analytical Ultracentrifugation: Sedimentation Velocity Analysis

### Abstract

The analytical ultracentrifuge is a high speed centrifuge with an optical system allowing observation of the concentration of macromolecules as a function of radius and time. In sedimentation velocity experiments,relatively high speeds are used so that a boundary is formed between the solution of sedimenting macromolecule and the buffer in which it is dissolved. Analysis of the rate boundary movement and evolution of its shape can yield information about the molar masses of species present as well as stoichiometries and equilibrium constants for their interactions. This overview discusses crucial issues pertaining to sedimentation velocity experiments, including non‐interacting and interacting systems, ideality and non‐ideality, and reversible versus kinetically limited equilibrium scenarios.

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### Literature Cited

Literature Cited | |

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Stafford, W.F. 1992. Boundary analysis in sedimentation transport experiments: A procedure for obtaining sedimentation coefficient distributions using the time derivative of the concentration profile. Anal. Biochem. 203:295‐301. | |

Stafford, W.F. 1994a. Boundary analysis in sedimentation velocity experiments. In Methods in Enzymology, Numerical Computer Methods, 240, part B (M.L. Johnson and L. Brandeds.) pp. 478‐501. Academic Press Orlando, Fla. | |

Stafford, W.F. 1994b. Sedimentation boundary analysis of interacting systems: Use of the apparent sedimentation coefficient distribution function. In Modern Analytical Ultracentrifugation: Acquisition and Interpretation of Data for Biological and Synthetic Polymer Systems (T.M. Schuster and T.M. Laueeds.) pp. 119‐137. Birkhäuser Boston Boston,Mass. | |

Stafford, W.F. 1998. Time difference sedimentation velocity analysis of rapidly reversible interacting systems: Determination of equilibrium constants by non‐linear curve fitting procedures. Biophys. J. 74:A301. | |

Stafford, W.F. and Schuster, T.M. 1995. Hydrodynamic methods. In Introduction to Biophysical Methods for Protein and Nucleic Acid Research (J.A. Glasel and M.P. Deutschereds.) pp. 111‐145. Academic Press, San Diego. | |

Stafford, W.F. and Sherwood, P. 2003. Analysis of heterologous interacting systems by sedimentation velocity: Curve‐fitting algorithms for estimation of sedimentation coefficients and equilibrium constants. J. Biophys. Chem. In press | |

Tanford, C. 1961. Physical Chemistry of Macromolecules. John Wiley & Sons, New York. | |

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Todd, G.P. and Haschmeyer, R.H. 1981. General solution to the inverse problem of the differential equation of the ultracentrifuge. Proc. Natl. Acad. Sci. U.S.A. 78:6739‐6743. | |

Van Holde, K.E. and Weischet, W.O. 1978. Boundary analysis of sedimentation‐velocity experiments with monodisperse and paucidisperse solutes. Biopolymers 17:1387‐1403. | |

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Williams, J.W., Van Holde, K.E., Baldwin, R.L., and Fujita, H. 1958. The theory of sedimentation analysis. Chem. Rev. 58:715‐806. | |

Key References | |

Williams et al., 1958. See above. | |

This selection of three pieces of older literature contains a tremendous amount of information that is very germane to modern day sedimentation analysis. | |

Schachman, 1959. See above. | |

Claesson and Moring‐Claesson, 1961. See above. |