Abstract
Many proteins function by changing conformation in response to ligand binding or changes in other factors in their environment. Any change in the sequence of a protein, for example during evolution, which alters the relative free energies of the different functional conformations changes the conditions under which the protein will function. Voltage-gated ion channels are membrane proteins that open and close an ion-selective pore in response to changes in transmembrane voltage. The charged S4 transmembrane helix transduces changes in transmembrane voltage into a change in protein internal energy by interacting with the rest of the channel protein through a combination of non-covalent interactions between adjacent helices and covalent interactions along the peptide backbone. However, the structural basis for the wide variation in the V50 value between different voltage-gated potassium channels is not well defined. To test the role of the loop linking the S3 helix and the S4 helix in voltage sensitivity, we have constructed a set of mutants of the rat Kv1.2 channel that vary solely in the length and composition of the extracellular loop that connects S4 to S3. We evaluated the effect of these different loop substitutions on the voltage sensitivity of the channel and compared these experimental results with molecular dynamics simulations of the loop structures. Here, we show that this loop has a significant role in setting the precise V50 of activation in Kv1 family channels.
Highlights
Potassium channels change conformation in response to transmembrane voltage
We evaluated the effect of these different loop substitutions on the voltage sensitivity of the channel and compared these experimental results with molecular dynamics simulations of the loop structures
We wanted to determine whether the distance between the ends of the helices, and the length of the linker, was the only constraint on the Gibbs free energy of activation (⌬Gopen) or if the loop was interacting with the rest of the channel to modulate V50
Summary
Potassium channels change conformation in response to transmembrane voltage. Results: The loop connecting the voltage sensing helix to an adjacent helix affects the voltage sensitivity. Voltage-gated ion channels are membrane proteins that open and close an ionselective pore in response to changes in transmembrane voltage. When the force on the S4 helix changes with membrane potential, the overall conformation of the channel protein changes The sum of these energetic interactions within the protein and between the protein and the lipid bilayer is responsible for setting the V50 of the channel, the voltage at which half of the channels are in an open state and half are in a closed state. The experimental results from the mutant channels were combined with molecular dynamics simulations of the same loops constrained by models of open and closed states of the channel We found that both the length and the composition of the loop are responsible for an energetic constraint on the transition between the opened and closed states. We conclude that the highly variable range of lengths and compositions of S3/S4 linkers found in nature is one factor in the quantitative evolution of voltage sensitivity in the Kv1 family of voltage-gated cation channels
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