2.A.4 The Cation Diffusion Facilitator (CDF) Family

The CDF family is a ubiquitous family, members of which are found in bacteria, archaea and eukaryotes (Paulsen and Saier, 1997). They transport heavy metals including cobalt, cadmium, zinc and possibly nickel, copper and mercuric ions. There are 10 mammalian paralogues (Cousins et al., 2006). Most members of the CDF family possess six putative transmembrane spanners with N- and C-termini on the cytoplasmic side of the membrane, but MSC2 of S. cerevisiae (TC #2.A.4.4.1) and Znt5 and hZTL1 of H. sapiens exhibit 15 and 12 putative TMSs, respectively (Cragg et al., 2002). These proteins exhibit an unusual degree of sequence divergence and size variation (300-750 residues). Eukaryotic proteins exhibit differences in cell localization. Some catalyze heavy metal uptake from the cytoplasm into various intracellular eukaryotic organelles (ZnT2-7) while others (ZnT1) catalyze efflux from the cytoplasm across the plasma membrane into the extracellular medium. Thus, some are found in plasma membranes while others are in organellar membranes such as vacuoles of plants and yeast and the golgi of animals (Chao and Fu, 2004b; Haney et al., 2005; MacDiarmid et al., 2003).

Prokaryotic and eukaryotic proteins cluster separately but probably function with the same polarity by similar mechanisms. These proteins are secondary carriers which utilize the pmf and function by H⁺ antiport (for metal efflux). One member, CzcD of Bacillus subtilis, has been shown to exchange the divalent cation (Zn²⁺ or Cd²⁺) for two monovalent cations (K⁺ and H⁺) in an electroneutral process energized by the transmembrane pH gradient (Guffanti et al., 2002). Another, ZitB of E. coli (TC #2.A.4.1.4), has been reconstituted in proteoliposomes and studied kinetically (Chao and Fu, 2004a). It appears to function by simple Me²⁺:H⁺ antiport with a 1:1 stoichiometry.

Montanini et al (2007) have conducted phylogenetic analysis of CDF family members. Their analysis revealed three major and two minor phylogenetic groups. They suggest that the three major groups segregated according to metal ion specificity: (1) Mn²⁺, (2) Fe²⁺ and Zn²⁺ as well as other metal ions, and (3) Zn²⁺ plus other metals, but not Iron.

At least two metal binding sites have been identified in the E. coli paralogue, YiiP (TC #2.A.4.1.5), and one plays a role in H⁺ binding as well (Chao and Fu, 2004b). The two Zn²⁺/Cd²⁺ binding sites consist of two interacting conserved aspartyl residues (Asp-157 and Asp-49), both in 2 fold symmetry-related TMS 5 and TMS 2, respectively, at the dimer interface of the homodimer (Wei and Fu, 2006). The (Asp-49–Asp157) may form a bimetal binding center. Two bound Cd²⁺ were transported cooperatively with sigmoidal dependency on the Cd²⁺ concentration. A translocation pathway for metal ions at the dimer interface has been proposed (Wei and Fu, 2006). CDF family members may generally be homodimeric (Haney et al., 2005; Wei et al., 2004).

Lu and Fu (2007) have reported the x-ray structure of YiiP of E. coli (2.A.4.1.5) in complex with zinc at 3.8 angstrom resolution. YiiP is a homodimer held together in a parallel orientation through four Zn²⁺ ions at the interface of the cytoplasmic domains.  The two transmembrane domains swing out to yield a Y-shaped structure. In each protomer, the cytoplasmic domain adopts a metallochaperone-like protein fold. The transmembrane domain features a bundle of six transmembrane helices and a tetrahedral Zn²⁺ binding site located in a cavity that is open to both the membrane outer leaflet and the periplasm. 

The generalized transport reaction for CDF family members is:

Me²⁺ (in) + H⁺ (out) ± K⁺ (out) Me²⁺ (out) + H⁺ (in) ± K⁺ (in).