Molecular Weight Determination of Protein-Dodecyl Sulfate Complexes by Gel Electrophoresis in a Discontinuous Buffer System

TLDR

The study reports on combining SDS gel electrophoresis with discontinuous buffer systems to analyze protein–SDS complexes. The method uses a sulfate‑borate discontinuous buffer to stack and separate protein–SDS complexes (2,300–320,000 Da) and evaluates them via Ferguson plots to determine retardation coefficients and apparent free mobilities. The analysis shows that apparent free mobilities are constant, yielding a linear log(relative mobility)–retardation coefficient relationship, and that the retardation coefficient is a uniform function of molecular weight, enabling molecular weight determination from negative log(relative mobility) versus molecular weight plots.

Abstract

This report describes methods and results obtained by combining the techniques of sodium dodecyl sulfate (SDS) gel electrophoresis and electrophoresis in discontinuous buffer systems. The SDS gel system utilizes a sulfate-borate discontinuity which stacks and unstacks protein-SDS complexes over a range of 2,300 to 320,000 daltons, providing high resolution fractionation. The properties of protein-SDS complexes are investigated by calculating retardation coefficients and apparent free mobilities from Ferguson plots. Apparent free mobilities are approximately constant, establishing a linear relationship between the logarithm of the relative mobility and the retardation coefficient. The retardation coefficient is shown both empirically and theoretically to be a uniform function of molecular weight of protein-SDS complexes over specified ranges, providing a rationale for determining molecular weight from plots of the negative logarithm of relative mobility against molecular weight.