Multiscale Kinetic Modeling of a Cl−/H+ Antiporter: Integrating Simulation and Experiment to Characterize A Complex Ion Exchange Process

Abstract

One of the central challenges in modern biophysics is understanding the molecular nature, and hence both the thermodynamics and kinetics, of coupled biomolecular transport mechanisms. The coupled ion exchange mechanism in Cl-/H+ antiporters offers an important and intriguing case study for this challenge. Herein, our new approach to multiscale kinetic modeling (MKM) and its application to characterize the Cl-/H+ exchange process in ClC-ec1 will be presented. We first calculate essential state-to-state rate coefficients with reactive and polarizable molecular dynamics simulations. A kinetic (Markov state) model is then used to optimize the rate coefficients within their calculated error to reproduce specific experimental rates and stoichiometry. This results in a set of solutions that not only predicts new, testable properties (e.g., rates at different pH values and the relative contribution of protein orientations), but also reveals insight into the series of transitions, the balance of rate-limiting steps, the molecular origin of the 2.2:1 Cl−/H+ stoichiometry, and the influence of protein orientation. Our results suggest that the consistent exchange ratio is a consequence of kinetic coupling wherein residue E148 plays an essential role through protonation-dependent anion transport, and an anion-dependent pKa value. They further demonstrate that a large conformational change is not essential at this level of modeling for the ion exchange mechanism, suggesting a more facile possible evolutionary connection to chloride channels. Finally, the results demonstrate how an ensemble of different exchange pathways, as opposed to a single series of transitions, culminates in the macroscopic observables and thereby explains the molecular mechanism.