TABLE OF CONTENTS Analytical ultracentrifugation is used to analyze protein self-association. It is employed to compare enzymatic activity and quaternary structure of proteins and to ascertain whether the crystal structure derived from X-ray scattering is identical with the solution structure. Furthermore, structure analysis by high-resolution NMR requires knowledge of the state of association and the type of equilibria. The classical approaches are the equilibrium sedimentation and the concentration-dependent analysis of weight-average sedimentation coefficients, the isotherm method. [1] Another possibility of the shape analysis of sedimentation velocity boundaries has been developed. [2] [4] These approaches provide association constants by fitting a set of sedimentation velocity profiles, which can be quickly obtained as an important aspect for less stable proteins. Furthermore, it is possible to omit the region near the cell base, which occasionally contain small quantities of aggregates. This is an advantage compared with the thermodynamically well-described method of sedimentation equilibrium. Our program allows the reliable determination of binding constants for self-associating macromolecules from sedimentation velocity runs.
[1]R. P. Frigon and S. N. Timasheff, Biochemistry, 1975, 14, 4559.
[2]J. -M. Claverie, H. Dreux and R. Cohen, Biopolymers, 1975, 14, 1685.
[4]P. Schuck, Anal. Biochem., 2003, 320, 104.
The program LAMMNUM with a numerical solution of the Lamm equation was written according to [2], [3] and [5]. It allows estimating equilibrium constants (K w). The program works with numerically calculated...
