Allowance for radial dilution in evaluating the concentration dependence of sedimentation coefficients for globular proteins

Abstract

The accuracy with which the concentration dependence of the sedimentation coefficient, s = s ⁰(1 − kc), can be quantified for globular proteins by commonly used procedures has been examined by subjecting simulated sedimentation velocity distributions for ovalbumin to c(s)‒s analysis. Because this procedure, as well as its g(s)‒s counterpart, is based on assumed constancy of s over the time course of sedimentation coefficient measurement in a given experiment, the best definition of the concentration coefficient k is obtained by associating the measured s with the mean of plateau concentrations for the initial and final distributions used for its determination. The return of a slightly underestimated k (by about 3%) is traced to minor mislocation of the air‒liquid meniscus position as the result of assuming time independence of s in a given experiment. Although more accurate quantification should result from later SEDFIT and SEDANAL programs incorporating the simultaneous evaluation of s ⁰ and k, the procedures based on assumed constancy of s suffice for determining the limiting sedimentation coefficient s ⁰—the objective of most s‒c dependence studies.

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