Abstract
Potassium channels are opened by ligands and/or membrane potential. In voltage-gated K+ channels and the prokaryotic KcsA channel, conduction is believed to result from opening of an intracellular constriction that prevents ion entry into the pore. On the other hand, numerous ligand-gated K+ channels lack such gate, suggesting that they may be activated by a change within the selectivity filter, a narrow region at the extracellular side of the pore. Using molecular dynamics simulations and electrophysiology measurements, we show that ligand-induced conformational changes in the KcsA channel removes steric restraints at the selectivity filter, thus resulting in structural fluctuations, reduced K+ affinity, and increased ion permeation. Such activation of the selectivity filter may be a universal gating mechanism within K+ channels. The occlusion of the pore at the level of the intracellular gate appears to be secondary.
Highlights
In voltage-gated potassium channels, the opening of an intracellular gate formed by a ‘bundle-crossing’ of the inner pore-lining helices is hypothesized to allow K ions to flow through the pore, down their electrochemical gradient (Yellen, 1998)
A narrow section of the pore located towards the extracellular side of the membrane, called the selectivity filter because it allows the pore to select for K ions, has been proposed to play double duty as an inactivation gate: following a stimulus, the pore opens at the intracellular bundle-crossing and, in the continued presence of the stimulus, the selectivity filter changes its conformation to prevent further flux of K ions
Our result did not depend on the initial ion configuration, as a potential of mean force (PMF) calculation started with K ions in sites S1-S3 and Cav with water molecules in S2 and S4, reveals similar free energy wells about 7 kcal/mol deep (Figure 1—figure supplement 1)
Summary
In voltage-gated potassium channels, the opening of an intracellular gate formed by a ‘bundle-crossing’ of the inner pore-lining helices is hypothesized to allow K ions to flow through the pore, down their electrochemical gradient (Yellen, 1998). Functional experiments examining state-dependent accessibility of pore blockers (Contreras and Holmgren, 2006; Wilkens and Aldrich, 2006; Rapedius et al, 2012; Posson et al, 2013b; Posson et al, 2015) or thiol-modifying agents (Flynn and Zagotta, 2001; Zhou et al, 2011), have shown that in other channels the intracellular pore-lining helices no longer form a bundle crossing that obstructs K+ flux in the closed state, strongly implicating the selectivity filter as an activation gate (Flynn and Zagotta, 2001; Proks et al, 2003; Bruening-Wright et al, 2007; Klein et al, 2007; Contreras et al, 2008) This suggests that different K+ channels with high sequence and structural homology do not share a universal gating mechanism. We challenge this statement based on our findings with KcsA, a structurally characterized model K+ channel, which was believed to activate by simple opening of the
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