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, Glu¹⁴⁸ and Tyr⁴⁴⁵, 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.