The problem with the Offord equation: Hydration - a statistical investigation of the effect of charge size and confirmation of the effect of charge distribution upon electrophoretic mobilities

A previous study has demonstrated that the average electrophoretic behaviou r (mobilities,,a,,) of eighteen peptides were well described by the inverse square law: mu (ep) alpha 1/r(2) (where r is the hydrodynamic radius). How ever, examination of the dependence of mu (ep) upon the magnitude of the ch arge and its distribution led to the conclusion that deviations from the av eraged behaviour arose from a charge-induced volumetric effect. In the current word we have sought to determine the generality of the above findings via a statistical examination of mu (ep) for an independent and l arger set of peptides. Both objective testing by the Offord eq. and statist ical analysis of the variation of the log mu (ep)/Z function reveals a stro ng residual dependence upon the magnitude of the overall charge. Furthermor e, it was found that an even finer distinction exists between peptides of d iffering charge distribution types. Terminal charges affect the peptide mob ilities differently to those charges located within the amino acid sequence and isolated charges affect mobility differently to the some number of adj acent charges. The empirical addition of hydration to each charge type prod uces a linear log plot of the Offord equation (R-2 = 0.991) that has a grad ient (0.64) very close to that theoretically expected (0.67). Hence hydrati on removes the observed deviations from the averaged electrophoretic behavi our that is associated with highly charged analytes and corrects the Offord equation. It is postulated that higher charge densities lead to more open structures, greater salvation and thus larger electromigrating molecular io ns with smaller mobilities.