A method for directly fitting the time derivative of sedimentation velocity data and an alternative algorithm for calculating sedimentation coefficient distribution functions.
The time-derivative method for deriving the sedimentation coefficient distribution, g(s*), from sedimentation velocity data that was developed by Walter Stafford has many advantages and is now widely used. By fitting Gaussian functions to the g(s*) distribution both sedimentation and diffusion coefficients (and therefore molecular masses) for individual species can be obtained. However, some of the approximations used in these procedures limit the accuracy of the results. An alternative approach is proposed in which the dc/dt data are fitted rather than g(s*). This new approach gives improved accuracy, extends the range to sedimentation coefficients below 1 S, and enhances resolution of multiple species. For both approaches the peaks from individual species are broadened when the data cover too wide a time span, and this effect is explored and quantified. An alternative algorithm for calculating ĝ(s*) from the dc/dt curves is presented and discussed. Rather than first averaging the dc/dt data for individual scan pairs and then calculating ĝ(s*) from that average, the ĝ(s*) distributions are calculated for every scan pair and then subsequently averaged. This alternative procedure yields smaller error bars for g(s*) and somewhat greater accuracy for fitted hydrodynamic properties when the time span becomes large.
Philo
J S
JS
Alliance Protein Laboratories, 3957 Corte Cancion, Thousand Oaks, California, 91360, USA.
D000465:Algorithms; D002499:Centrifugation, Density Gradient / Q000706:statistics & numerical data*; D003627:Data Interpretation, Statistical; D005069:Evaluation Studies as Topic