Characterization of the ZAT1p zinc transporter from Arabidopsis thaliana in microbial model organisms and reconstituted proteoliposomes

The ZAT1p zinc transporter from Arabidopsis thaliana (L.) Heynh. is a member of the cation diffusion facilitator (CDF) protein family. When heterologously expressed in Escherichia coli, ZAT1p bound zinc in a metal blot. Binding of zinc occurred mainly to the hydrophilic amino acid region from H-182 to H-232. A ZAT1p/ZAT1p*Delta(M-1-I-25) protein mixture was purified and reconstituted into proteoliposomes. Uptake of zinc into the proteoliposomes did not require a proton gradient across the liposomal membrane. ZAT1p did not transport cobalt, and transported cadmium at only 1% of the zinc transport rate. ZAT1p functioned as an uptake system for Zn-65(2+) in two strains of the Gram-negative bacterium Ralstonia metallidurans, which were different in their content of zinc-efflux systems. The ZAT1 gene did not rescue increased zinc sensitivity of a DeltaZRC1 single-mutant strain or of a DeltaZRC1 DeltaCOT1 double-mutant strain of Saccharomyces cerevisiae, but ZAT1 complemented this phenotype in a DeltaSpZRC1 mutant strain of Schizosaccharomyces pombe.