Determination of thecis-trans isomerization barrier of severalL-peptidyl-L-proline dipeptides by dynamic capillary electrophoresis and computer simulation

Abstract

Dynamic capillary electrophoresis (DCE) and computer simulation of the elution profiles with the theoretical plate and the stochastic model has been applied to determine the isomerization barriers of the three dipeptides L-alanyl-L-proline, L-leucyl-L-proline, and L-phenylalanyl-L-proline. The separation of the rotational cis-trans isomers has been performed in an aqueous 70 mM borate buffer at pH 9.5. Interconversion profiles featuring plateau formation and peak broadening were observed. To determine the rate constants k1 and k(-1) of the cis-trans isomerization in dynamic capillary electrophoresis, equations have been derived for the theoretical plate model and stochastic model. The electropherograms were simulated with the ChromWin software which uses the experimental data plateau height h(plateau), peak width at half height Wh, the total migration times of the cis-trans isomers tR and the electroosmotic break-through time t0 as well as the peak ratio [cis]/[trans]. From temperature-dependent measurements, the rate constants k1 and k(-1) and the kinetic activation parameters deltaG#, deltaH# and deltaS# of the cis-trans isomerization of the three dipeptides were obtained.

Separate Determination of Mobile‐Phase Rate Constants for Reversible Reactions

“…The mobile phase rate constant can be evaluated by plotting the reduced rate constant k E t E vs. retention times of the reactants, if only the residence time The rate constants evaluated for the cis -trans isomerization of Leu-Pro are of the same magnitude as the rate constants published by Schoetz et al [8] from capillary electrophoresis. The remaining differences will be due to different solvents, pH, and buffers.…”

“…[8] In the RP liquid chromatography, it was found that the reaction in the stationary phase is considerably faster than in the mobile phase.…”

The Reciprocal Principle of Selectand-Selector-Systems in Supramolecular Chromatography †

“…The reciprocal concept of interconverting molecules in separation methods could also be extended to the liquid chromatographic investigation of conformational cis/trans -diastereomers of l -peptidyl- l -proline. Studies of proline-containing peptides separated on calixarene-bonded silica gels have been investigated by Gebauer et al [131] and interconversion barriers of l -proline containing dipeptides by dynamic CE were determined [132].…”

Native state dynamics affects the folding transition of porcine pancreatic phospholipase A2

“…Though no global unfolding seems to be required to expose either of the identified cleavage sites, the native state of Ca 2 free ppPLA 2 at pH 8.0 is clearly highly dynamic. As evidenced by their activation free energy barriers, which are distinct, but not high ( Table 2), those of peptidyl-proline cis/trans isomerization being about one third higher [70][71][72], these opening reactions are minor in scale and can proceed readily at native conditions.…”