Charge Reversion to Charge Carrying Positions of S4 in Voltage Gated Shaker K-Channels

Abstract

Listen

Translate article

Voltage gated Shaker potassium channels increases open probability (Po) by 20-fold with a ∼6 mV depolarization. Such high voltage sensitivity is mostly due to the electrophoretic transmembrane relocation of four arginines residues in each of their four voltage sensing protein domains (VSD). These arginines movement across the electric field make possible channel opening upon membrane depolarization. We tested if the positions occupied by the voltage sensing arginines could carry acidic residues. We mutated three of these positions to aspartate: Arg362, Arg365, and Arg368 on an N-type inactivation removed background Shaker channel. All mutations were introduced with the use of the QuikChange kit and the mutation was verified by sequencing. Heterologous expressed in Xenopus oocytes, all mutants showed levels of expression comparable to that of the native channel. To determine voltage sensitivity of these charge reverting modified channels, we measured the voltage dependency of Po at voltages negative enough to observe only sporadic single channel openings in membrane patches containing hundreds of channels. From the exponential relation between Po and voltage we estimated the effective valence of opening in the range of Po ∼10-6. All charge mutants showed an effective valence ∼50% of that of the native Shaker. These results together with the comparable level of channel expression in oocytes are consistent with the idea the voltage sensitive positions in S4 are not specific for basic residues.VGP was a doctoral Conicyt fellow and DN was financed by Fondecyt #1090493