Abstract
The gating of voltage-gated ion channels is controlled by the arginine-rich S4 helix of the voltage-sensor domain moving in response to an external potential. Recent studies have suggested that S4 moves in three to four steps to open the conducting pore, thus visiting several intermediate conformations during gating. However, the exact conformational changes are not known in detail. For instance, it has been suggested that there is a local rotation in the helix corresponding to short segments of a 3-helix moving along S4 during opening and closing. Here, we have explored the energetics of the transition between the fully open state (based on the X-ray structure) and the first intermediate state towards channel closing (C), modeled from experimental constraints. We show that conformations within 3 Å of the X-ray structure are obtained in simulations starting from the C model, and directly observe the previously suggested sliding 3-helix region in S4. Through systematic free energy calculations, we show that the C state is a stable intermediate conformation and determine free energy profiles for moving between the states without constraints. Mutations indicate several residues in a narrow hydrophobic band in the voltage sensor contribute to the barrier between the open and C states, with F233 in the S2 helix having the largest influence. Substitution for smaller amino acids reduces the transition cost, while introduction of a larger ring increases it, largely confirming experimental activation shift results. There is a systematic correlation between the local aromatic ring rotation, the arginine barrier crossing, and the corresponding relative free energy. In particular, it appears to be more advantageous for the F233 side chain to rotate towards the extracellular side when arginines cross the hydrophobic region.
Highlights
Voltage-gated ion channels are membrane proteins that conduct ions, regulated by the electrostatic potential across the membrane
The present study shows that our recent models of the intermediate C1 state from experiments is a metastable conformation with a local free energy minimum, and it appears to directly confirm a 310-helix region that slides along the S4 sequence as the helix moves from one intermediate conformation to the
Rather than blindly moving between unknown states, we have shown the end conformation of the present steered molecular dynamics simulations to be remarkably close to the Xray structure of the chimera voltage sensor, both in terms of sidechain interactions, 310-helix contents, and not least a 3 A RMSD
Summary
Voltage-gated ion channels are membrane proteins that conduct ions, regulated by the electrostatic potential across the membrane These channels play fundamental roles for instance in the generation and propagation of nerve impulses and in cell homeostasis. They are made up of four homologous domains, each of which contains six transmembrane helices. As the electrostatic potential across the membrane is changed, these charges will be subject to large forces that cause S4 to move in the voltage sensor This in turn initiates a conformation change in the pore domain that opens or closes the channel to control the flow of Kz ions from the intracellular to the extracellular side [1,2,3,4,5,6]
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