The CLC-family protein CLC-ec1, a bacterial homologue of known structure, stoichiometrically exchanges two Cl − for one H + via an unknown membrane transport mechanism. This study examines mutations at a conserved tyrosine residue, Y445, that directly coordinates a Cl − ion located near the center of the membrane. Mutations at this position lead to “uncoupling,” such that the H + /Cl − transport ratio decreases roughly with the volume of the substituted side chain. The uncoupled proteins are still able to pump protons uphill when driven by a Cl − gradient, but the extent and rate of this H + pumping is weaker in the more uncoupled variants. Uncoupling is accompanied by conductive Cl − transport that is not linked to counter-movement of H + , i.e., a “leak.” The unitary Cl − transport rate, measured in reconstituted liposomes by both a conventional initial-velocity method and a novel Poisson dilution approach, is ∼4,000 s −1 for wild-type protein, and the uncoupled mutants transport Cl − at similar rates.