Rigorous estimation of effective protein charge from experimental electrophoretic mobilities for proteomics analysis using microchip electrophoresis

Understanding of electrophoretic mobility and zeta potential is an important issue in charge characteristics of proteins for proteomics research. By exploring the electrostatic and electrokinetic principles, we presented a framework that would allow rigorous interpretations of surface zeta potential and effective charge for highly charged conditions holding the full range of buffer pH. Together with an analytic expression by considering the Henryʹs formula with Debye–Hückel ansatz, the zeta potential at the protein surface was evaluated from an integral expression for the electrophoretic mobility of a spherical particle on the basis of nonlinear Poisson–Boltzmann equation. Subsequently, the effective protein charge was evaluated from the corresponding relationships between potential distribution and surface charge, with accounting for the presence of an electric double layer. Applying the microchip electrophoresis, the experimental results of mobility data were obtained for model proteins with bovine serum albumin and ovalbumin in different pH of buffer solution. To illustrate the usefulness of our considerations, experimental data of electrophoretic mobility available in the literature are also included. It is evident that the results by linear correlations are identified to overestimate the zeta potential, while they underestimate the effective charge. The discrepancy between linear and nonlinear correlations is trivial for the zeta potential less than 25.69 mV, however, it fairly increases with increasing the absolute value of zeta potential.