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Biophys J, March 2000, p. 1606-1619, Vol. 78, No. 3
Molecular Interactions Resource, Bioengineering and Physical Science Program, ORS, National Institutes of Health, Bethesda, Maryland 20892 USA
A new method for the size-distribution analysis of
polymers by sedimentation velocity analytical ultracentrifugation is
described. It exploits the ability of Lamm equation modeling to
discriminate between the spreading of the sedimentation boundary
arising from sample heterogeneity and from diffusion. Finite element
solutions of the Lamm equation for a large number of discrete
noninteracting species are combined with maximum entropy regularization
to represent a continuous size-distribution. As in the program CONTIN,
the parameter governing the regularization constraint is adjusted by
variance analysis to a predefined confidence level. Estimates of the
partial specific volume and the frictional ratio of the macromolecules
are used to calculate the diffusion coefficients, resulting in
relatively high-resolution sedimentation coefficient distributions
c(s) or molar mass distributions
c(M). It can be applied to interference
optical data that exhibit systematic noise components, and it does not
require solution or solvent plateaus to be established. More details on
the size-distribution can be obtained than from van Holde-Weischet
analysis. The sensitivity to the values of the regularization parameter
and to the shape parameters is explored with the help of simulated
sedimentation data of discrete and continuous model size distributions,
and by applications to experimental data of continuous and discrete protein mixtures.
Biophys J, March 2000, p. 1606-1619, Vol. 78, No. 3
© 2000 by the Biophysical Society 0006-3495/00/03/1606/14 $2.00
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