Figure 5.

Characterization of Zn²⁺ binding to the first extracellular loop of mZIP4. A, WT (peptide 1) and mutant (peptide 2) peptide sequences corresponding to the first extracellular loop region used in the zinc-binding assay. B–D, zinc binding analysis of peptides 1 and 2. The elution profiles are shown from a PD-10 column of 100 nmol of total zinc ions alone (B), 100 nmol of total zinc ions with 1.0 molar eq of peptide 1 (C), 100 nmol of zinc with 1.0 molar eq of peptide 2 (D). Concentrations of peptide are shown as filled circles with dashed lines, and eluted zinc levels are shown as open squares with a solid line. E, the stoichiometry of zinc binding under varied concentrations of the peptide 1 and zinc ions. F, zinc facilitates cross-linking of peptide 1 but not peptide 2. Peptides 1 and 2 (0.1 mm) were treated with 0.5 mm ZnCl₂ and/or 2.5 mm of chemical cross-linker DSS and resolved on a Tris Tricine gel after quenching with 50 mm Tris (pH 7.5).