Charge reversal supports an electrostatic mechanism for complementation. Currents were studied as in Fig. 2 A. Bar graphs plot current amplitudes ± SEM at −100 mV. (A) S104E ΔK_2PØ with no other change, F137 in TM2, or V258 in TM4 altered to lysine (K) or arginine (R). (insets) Glutamate (E) in P1 (red); TM sites studied (blue). Statistical differences versus S104E are indicated (n = 10). (B) WT ΔK_2PØ or ΔK_2PØ with single changes at position F137 (green box). Statistical differences versus WT are indicated (n = 6–10). Mutation of F137 increases single-channel activity (i.e., WT, open probability = ∼0.77; F137C, open probability = ∼0.93) and thereby decreases closed state-dependent Zn²⁺ block (i.e., WT, half-maximal inhibition at equilibrium (K_i) = 5 μM; F137A, K, and D, K_i >> 1 mM) by reported methods (Zilberberg et al., 2001). (C) S104K ΔK_2PØ channels with no other change or second site changes at F137. (inset) S104K in P1 (blue); TM2 site studied (red). Statistical differences versus S104K are indicated (n = 4–10). (D) S104E ΔK_2PØ channels with no other change or second site changes at F137. (inset) S104E in P1 (red); TM2 site studied (blue). Statistical differences versus S104E are indicated (n = 6–8). (E) Sample current traces for the indicated ΔK_2PØ channels with one or two mutations. Bars, 5 μA and 100 ms. *, P < 0.05; **, P < 0.01; ***, P < 0.001.