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

Göran Klement ^*, Johanna Nilsson ^*, Peter Århem ^* and Fredrik Elinder 

^* The Nobel Institute for Neurophysiology, Department of Neuroscience, Karolinska Institutet, SE-171 77 Stockholm, Sweden; and Department of Clinical and Experimental Medicine, Division of Cell Biology, Linköping University, SE-581 85 Linköping, Sweden

Correspondence: Address reprint requests to Peter Århem, The Nobel Institute for Neurophysiology, Dept. of Neuroscience, Karolinska Institutet, SE-171 77 Stockholm, Sweden. Tel.: 46-8-524-869-03; E-mail: peter.arhem{at}ki.se.

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 (₁ = 20 ms, ₂ = 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 ( >> 10 min), whereas positive voltage steps quickly do ( = 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.