Modulating the Chemical Transport Properties of the CLC Antiporter via Alternative Anion Flux and Mutation

Abstract

ClC-ec1, a prototype of the ClC family of proteins which are ubiquitous in nature and govern a wide range of physiological processes, enables the transmembrane exchange of Cl− for H+ in opposite directions. It has been known that other polyatomic anions, such as NO3− and SCN−, are transported by ClC-ec1, but proton flux is partly or completely uncoupled with anions fluxes. Herein, with the help of multiscale simulations in which the Grotthuss mechanism of proton transport (PT) is treated explicitly, we report, to our knowledge, the first free energy profiles for PT and coupled change of the water density in the channel with either NO3− or SCN− present at the central site. We also discovered how a newly formed phenylalanine gate can interrupt the water structure in the I109F mutant. The proton conductance calculated from the free energy profile recaptures previous experiment results showing reduced or abolished proton flux in polyatomic anion-bound wild-type ClC, compared to that coupled with Cl− flux. By analyzing the free energy profile, we have discovered that most relevant to the PT barrier is the water connectivity along the PT pathway in the presence of the excess proton, instead of the initial hydration of the cavity. The water structure can be modulated by the property of the anion or the gate formed by the residues. The results presented here suggest how water environment and PT are mutually affected as well as how we should control the functionality of the ClC family of proteins.