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

摘要

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.