Peptide mapping by capillary zone electrophoresis: How close is theoretical simulation to experimental determination

A multi-variable computer model is presented for the prediction of the elec trophoretic mobilities of peptides at pH 2.5 from known physico-chemical co nstants of their amino acid residues. The model is empirical and does not c laim any theoretical dependencies; however, the results suggest that, at le ast at this pH, peptides may be theoretically represented as classical poly mers of freely joined amino acid residues of unequal sizes. The model assum es that the electrophoretic mobility can be represented by a product of thr ee functions that return the contributions of peptide charge, length and wi dth, respectively to the mobility. The model relies on accurate experimenta l determination of the electrophoretic mobilities of a diverse set of pepti des, by capillary zone electrophoresis (CZE), at 22 degreesC, with a 50 mM phosphate buffer, at pH 2.5. The electrophoretic mobilities of a basis set of 102 peptides that varied in charge from 0.65 to 16 and in size from two to 42 amino acid residues were accurately measured at these fixed experimen tal conditions using a stable 10% linear polyacrylamide-coated column. Data from this basis set was used to derive the peptide charge, length, and wid th functions respectively. The main purpose of this endeavor is to use the model for the prediction of peptide mobilities at pH 2.5, and for simulatio n of CZE peptide maps of protein digests. Excellent agreement was obtained between predicted and experimental electrophoretic mobilities for all categ ories of peptides, including the highly charged and the hydrophobic. To ill ustrate the utility of this model in protein studies it was used to simulat e theoretical peptide maps of the digests of glucagon and horse cytochrome c. The resulting maps were compared and contrasted with their experimental counterparts. The potential of this approach and its limitations are discus sed. Published by Elsevier Science B.V.