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

Abstract

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.