Ion permeation through a Cl--selective channel designed from a CLC Cl-/H+ exchanger

The CLC family of Cl--transporting proteins includes both Cl- channels and Cl-/H+ exchange transporters. CLC-ec1, a structurally known bacterial homolog of the transporter subclass, exchanges two Cl- ions per proton with strict, obligatory stoichiometry. Point mutations at two residues, Glu(148) and Tyr(445), are known to impair H+ movement while preserving Cl- transport. In the x-ray crystal structure of CLC-ec1, these residues form putative gates flanking an ion-binding region. In mutants with both of the gate-forming side chains reduced in size, H+ transport is abolished, and unitary Cl- transport rates are greatly increased, well above values expected for transporter mechanisms. Cl- transport rates increase as side-chain volume at these positions is decreased. The crystal structure of a doubly ungated mutant shows a narrow conduit traversing the entire protein transmembrane width. These characteristics suggest that Cl- flux through uncoupled, ungated CLC-ec1 occurs via a channel-like electrodiffusion mechanism rather than an alternating-exposure conformational cycle that has been rendered proton-independent by the gate mutations.