Voltage dependence of gating charge movement and gating pore opening of R2C measured from a holding potential of 0 mV. (A) Example of gating currents measured in a cell expressing R2C in response to hyperpolarizations from a holding potential of 0 mV. (B) R2C gating pore currents elicited by holding the cell at 0 mV and applying a series of hyperpolarizing pulses in 10-mV increments with no leak subtraction. I_gp was measured at the end of the pulse. (C) The correlation of charge movement (open squares; Q-V: V_1/2 = −115 ± 3 mV, k = 23.7 ± 0.6 mV; n = 6) and gating pore conductance (blue circles; G_gp: V_1/2 = −139.3 ± 3.2 mV, k = −21.6 ± 2.6 mV; n = 5) of NaChBac R2C mutant. The Q-V (black dashes) and G_gp-V (red dashes) relationships for R1C from Fig. 2 C are overlain for comparison. Error bars represent SEM. (D) A cartoon illustrating our interpretation of the results for R2C. R3 occupies and seals the gating pore at 0 mV. By hyperpolarizing to more negative potentials, S4 moves inward, bringing R2C into the critical position in the gating pore negative to approximately −90 mV.