The Cu²⁺ coordination sites are conserved in Shaker channels. (A) I_K evoked from WT Shaker channels in response to pulses to −10 mV from a holding potential of −90 mV at the indicated [Cu²⁺] (0–100 μM). (B) G_K-V relations from the same patch in different [Cu²⁺] are normalized by G_Kmax in 0 Cu²⁺. (0 (○), 1 (•), 5 (▾), 20 (▴), and 100 μM Cu²⁺ (▪)). (C) I_K at 0 mV from WT channels at the indicated [Zn²⁺] (0–10,000 μM). (D) G_K-V relations in different [Zn²⁺], normalized by G_Kmax in 0 Zn²⁺ (0 (○), 100 (•), 1,000 (▾), 4,000 (▴), 10,000 μM Zn²⁺ ()). (E) I_K at 0 mV from E247A channels in 0, 1, and 5 μM Cu²⁺. (F) I_K at +30 mV from E247C in 0, 0.1, 1, and 5 μM Cu²⁺. (G and H) I_K at +10 mV from E247A (G) or E247C (H) in 0, 100, and 1,000 μM Zn²⁺. (I and J) I_K from R365A at 0 mV (I) or R365C at −40 mV(J) in 0, 1, and 5 μM Cu²⁺. All Shaker experiments were performed from a holding potential of −90 mV. Most data were obtained in standard Shaker internal and external solutions (see Materials and methods) with the exception of R365 mutant data in I and J, which were obtained in a low chloride external solution with all but 10 mM Cl⁻ replaced by MeO₃⁻, as in standard mSlo1 external solution.