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pubmed:abstractText |
Sequence homology of the Escherichia coli YiiP places it within the family of cation diffusion facilitators, a family of membrane transporters that play a central role in regulating cellular zinc homeostasis. Here we describe the first thermodynamic and mechanistic studies of metal binding to a cation diffusion facilitator. Isothermal titration calorimetric analyses of the purified YiiP and binding competitions among Zn(2+), Cd(2+), and Hg(2+) revealed a mutually competitive binding site common to three metal ions and a set of noncompetitive binding sites, including one Cd(2+) site, one Hg(2+) site, and at least one Zn(2+) site, to which the binding of Zn(2+) exhibited partial inhibitions of both Cd(2+) and Hg(2+) bindings. Lowering the pH from 7.0 to 5.5 inhibited binding of Zn(2+) and Cd(2+) to the common site. Further, the enthalpy change of the Cd(2+) binding to the common site was found to be related linearly to the ionization enthalpy of the pH buffer with a slope corresponding to the release of 1.23 H(+) for each Cd(2+) binding. These H(+) effects are consistent with a coupled deprotonation process upon binding of Zn(2+) and Cd(2+). Modification of histidine residues by diethyl pyrocarbonate specifically inhibited Zn(2+) binding to the common binding site, indicating that the mechanism of binding-deprotonation coupling involves a histidine residue(s).
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