A Tyrosine Substitution in the Cavity Wall of a K Channel Induces an Inverted Inactivation

Ion permeation and gating kinetics of voltage-gated K channels critically depend on the amino-acid composition of the cavity wall. Residue 470 in the Shaker K channel is an isoleucine, making the cavity volume in a closed channel insufficiently large for a hydrated K+ ion. In the cardiac human ether-a-go-go-related gene channel, which exhibits slow activation and fast inactivation, the corresponding residue is tyrosine. To explore the role of a tyrosine at this position in the Shaker channel, we studied I470Y. The activation became slower, and the inactivation faster and more complex. At +60 mV the channel inactivated with two distinct rates (τ1 = 20 ms, τ2 = 400 ms). Experiments with tetraethylammonium and high K+ concentrations suggest that the slower component was of the P/C-type. In addition, an inactivation component with inverted voltage dependence was introduced. A step to −40 mV inactivates the channel with a time constant of 500 ms. Negative voltage steps do not cause the channel to recover from this inactivated state (τ ≫ 10 min), whereas positive voltage steps quickly do (τ = 2 ms at +60 mV). The experimental findings can be explained by a simple branched kinetic model with two inactivation pathways from the open state.