Figure 3: Y578 mutations affect Zn²⁺ inhibition of common gating.

(a) Relationship between Zn²⁺-binding residues C277 and C278, residue F235, and residues E232 (E_ext) and Y578 (Y_cen) in the inactivated ClC-1 homology model, based on EcClC coordinates. In these panels helix G is coloured green and other helices and residues are coloured as in Fig. 1 (helix F, blue; helix R, yellow). Similarly, anion-binding sites are depicted as green spheres, and are labelled S_ext (e) and S_int (i). (b) Inhibition of WT ClC-1, F235L and Y578 mutant channels by 1 mM extracellular Zn²⁺. Data points represent peak current amplitude elicited by a 10 ms pulse to −100 mV, applied from a holding potential of −30 mV at 4-s intervals (inset), normalized to current amplitude preceding application of Zn²⁺. Data are means from WT n=5, F235L n=4, Y578A n=3, Y578F n=4–5, Y578E n=3, Y578H n=4–12 and Y578K n=3 (separate experiments). Dotted lines represent s.e.m. Dashed lines representing the fit of a single exponential function with an added minimum asymptote to WT ClC-1 data (τ=378±17 s, minimum asymptote=0.08±0.02) are included in each panel for the purpose of comparison. Representative current traces and gating curves for mutant F235L are shown in Supplementary Fig. S2.