A661C and I663C primarily affect activation time course. (A) Conductance versus voltage relations for A661C and I663C. (B) Current traces of slowly activating mutant channels A661C and I663C. Traces were evoked by a depolarizing pulse to +40 mV from a holding potential of −80 mV. Dashed line indicates zero current level. (C) Time constants of activation over the range of voltages tested. Activation traces as in B were fit with a single exponential function, ignoring initial sigmoidicity so as to capture the kinetics of the final closed to open transition. n = 8–10 for each data point. The activation time constant is voltage dependent and slower over the range of voltages. (D) Cross-sectional membrane view showing A661 (orange) and I663 (green) side chains based on the closed-state MlotiK1 homology model. (E) View from the cytoplasm of the closed-state MlotiK1 homology model, illustrating the proximity of A661 and I663 side chains from neighboring subunits. (F) Close-up view shows A661 and I663 side chain interaction in the closed state. (G) Cross-sectional membrane view of A661 and I663 in the open-state rKv1.2 homology model. (H) View of A661 and I663 from the cytoplasm in the open state. (I) Close-up view of A661 and I663 in open state.