15] Structure—Mobility relationships in free solution zone electrophoresis

Abstract

Publisher Summary The correlation of the structure of a solute, with electrophoretic mobility, is of interest for numerous reasons. Mobility in free solution gives a direct measure of the net effective charge of the solute that in turn can be a sensitive and direct measure of many solution phase interactions. Among these are stability, activity, and solubility that for solutes, such as proteins, the subject of this work, are all influenced by solution pH and the charge state of the protein. From a bioanalytical perspective, a knowledge of how solute structure affects mobility aids in the optimization of high-resolution separations. In general, correlation of solute structure, with mobility or the equivalent charge state of a solute, is fundamental to an understanding of how proteins behave in solution. With this in mind, it is noteworthy that capillary electrophoresis (CE) conducted in free solution is an ideal means of determining the effective net charge of proteins. This chapter discusses the results of investigations that have shown that mobility predictions of peptides and proteins, as a function of solution pH, are attainable if the amino acid content of the solute is known. As an introduction to this subject, the body of this chapter discusses that the easily calculated valence-pH titration curves for proteins are directly proportional to mobility-pH titration curves. The appendix at the end of this chapter extends the discussion, emphasizing calculated and actual protein valence and demonstrating how equations derived by inspection in the body of the work are arrived at fundamentally.