348C of TM6 forms a tight Cd²⁺-binding site with either 1144C or 1145C of TM12. (A) Acceleration of current recovery from Cd²⁺-induced inhibition of 348C/1144C currents by DTT in an inside-out recording. Although 348C/1144C currents could recover from the Cd²⁺-inhibited state irrespective of the presence of DTT, comparison of the time constants generated by fitting both current relaxations with a single exponential function (red and blue solid curves) indicates DTT can accelerate such a process. (B) Comparison of 348C/1144C current relaxation time constants indicates there is significant difference when DTT is in the solution. **, P < 0.01. (C) DTT-dependent recovery of 348C/1145C inside-out current after Cd²⁺ application implicates a tight Cd²⁺-binding site constructed by these two cysteines. (D) Summary of effect of Cd²⁺ on each mutant as indicated in the panel. Mean ± SEM is shown.