Voltage Sensor Immobilization in Kv1.2 Channels

Abstract

The voltage gated potassium channel family contains several functionally distinct isoforms which serve to shape the duration, frequency and timing of action potential firing in electrically excitable cells. Several mechanistic elements that contribute to the function of the voltage sensing apparatus are not yet fully described, especially in channels other than the prototypical Shaker potassium channel. We have studied voltage sensor rearrangements of Kv1.2 channels using two-electrode voltage clamp fluorometry and gating current recordings in mammalian cells. Fluorescence measurements reporting on voltage sensor movement revealed a transition into a relaxed state upon prolonged depolarization, causing a left shift in the fluorescence-voltage relationship. Gating current measurements of wild type Kv1.2 channels recorded in permeant ion free solutions (NMDG+int//TEA+ext) also displayed a left shifted Q-V after depolarizing pre-pulses, reflecting a stabilization of the activated state of the voltage sensor. To enable examination of the effects of different cations and processes of inactivation in Kv1.2 a non-conducting double mutant channel mimicking the permanently slow inactivated and non-conducting Shaker W434F, Kv1.2(W366F, V381T) was created. Off-gating currents recorded from this channel in the presence of permeant ions displayed less voltage sensor stabilization in the activated state than the wild type channel. These data suggest that cations play a specific role in regulating voltage sensor dynamics in the Kv1.2 channel.