Biophysical characterization of membrane and cytoskeletal proteins by sedimentation analysis.

Abstract

Analytical ultracentrifugation can be used to determine a number of important properties of proteins including molecular weight, the thermodynamic parameters governing self-associations and heterogeneous interactions, and nonideality. Hydrodynamic properties including the sedimentation, diffusion, and factional coefficients, and molecular shape can also be determined. There are three fundamental experiments from which information can be obtained: (1) sedimentation velocity, in which the rate of transport of a sedimenting protein boundary is measured, (2) sedimentation equilibrium, in which the concentration distribution of protein is measured in the absence of net flow, and (3) diffusion experiments, in which the rate of spreading of a protein boundary is determined. Usually the data are obtained using one or more of the optical systems available on an analytical ultracentrifuge (Schachman, 1959; Svedberg and Pederson, 1940; Van Holde, 1971). However, the preparative ultracentrifuge, in combination with precise fractionation techniques, can provide information of similar accuracy in many cases (Attri and Minton, 1983, 1984, 1986; Howlett, 1987; Minton, 1989).