Fluoride-dependent interruption of the transport cycle of a CLC Cl(−)/H(+) antiporter

Cl(−)/H(+) antiporters of the CLC superfamily transport anions across biological membranes in varied physiological contexts. These proteins are weakly selective among anions commonly studied, including Cl(−), Br(−), I(−,)NO(3)(−), and SCN(−), but appear to be very selective against F(−). The recent discovery of a new CLC clade of F(−)/H(+) antiporters, which are highly selective for F(−) over Cl(−), led us to investigate the mechanism of Cl(−)-over-F(−) selectivity by a CLC Cl(−)/H(+) antiporter, CLC-ec1. By subjecting purified CLC-ec1 to anion transport measurements, electrophysiological recording, equilibrium ligand-binding studies, and x-ray crystallography, we show that F(−) binds in the Cl(−) transport pathway with affinity similar to Cl(−), but stalls the transport cycle. Examination of various mutant antiporters implies a “lock-down” mechanism of F(−) inhibition, in which F(−), by virtue of its unique H-bonding chemistry, greatly retards a proton-linked conformational change essential for the transport cycle of CLC-ec1.