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Many voltage-gated potassium (Kv) channels display a time-dependent phenomenon called C-type inactivation, whereby prolonged activation by voltage leads to the inhibition of ionic conduction, a process that involves a conformational change at the selectivity filter toward a non-conductive state. Recently, a high-resolution structure of a strongly inactivated triple-mutant channel kv1.2-kv2.1-3m revealed a novel conformation of the selectivity filter that is dilated at its outer end, distinct from the well-characterized conductive state. While the experimental structure was interpreted as the elusive non-conductive state, our molecular dynamics simulations and electrophysiological measurements show that the dilated filter of kv1.2-kv2.1-3m is conductive and, as such, cannot completely account for the inactivation of the channel observed in the structural experiments. The simulation shows that an additional conformational change, implicating isoleucine residues at position 398 along the pore lining segment S6, is required to effectively block ion conduction. The I398 residues from the four subunits act as a state-dependent hydrophobic gate located immediately beneath the selectivity filter. These observations are corroborated by electrophysiological experiments showing that ion permeation can be resumed in the kv1.2-kv2.1-3m channel when I398 is mutated to an asparagine—a mutation that does not abolish C-type inactivation since digitoxin (AgTxII) fails to block the ionic permeation of kv1.2-kv2.1-3m_I398N. As a critical piece of the C-type inactivation machinery, this structural feature is the potential target of a broad class of quaternary ammonium (QA) blockers and negatively charged activators thus opening new research directions toward the development of drugs that specifically modulate gating states of Kv channels.
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A sodium channel mutant removes fast inactivation with the inactivation particle bound
Fast inactivation is a key feature of voltage-gated sodium channels and is pivotal for countless physiological functions. Despite the prevalence of the canonical ball-and-chain model, more recent structural results suggest that fast inactivation requires multiple conformational changes beyond the binding of the inactivation particle, the IFM motif. Combining ionic current, gating current, and fluorescent measurements, here we showed that a double mutant at the bottom of the pore domain (CW) removes fast inactivation by interrupting the communication of the IFM motif and the pore. Instead of triggering fast inactivation, the IFM motif binding in CW allows the channel to enter an alternative open state. This alternative open state severely influenced the voltage sensor movements and was not accessible to wild type or other fast inactivation–deficient channels. Our results highlight the multistep nature of the fast inactivation process in mammalian voltage-gated sodium channels and demonstrate that CW modifies the channel behaviors more profoundly than simple removal of fast inactivation.
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- Award ID(s):
- 2121044
- PAR ID:
- 10557135
- Publisher / Repository:
- DOI PREFIX: 10.1085
- Date Published:
- Journal Name:
- Journal of General Physiology
- Volume:
- 157
- Issue:
- 1
- ISSN:
- 0022-1295
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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