Ion conduction and conformational flexibility of a bacterial voltage-gated sodium channel

Significance Voltage-gated sodium channels are one of the most fundamental electrical components in the nervous system and are key targets for local anesthesia and therapeutics for neurological and cardiac disorders. We have used multimicrosecond simulations to provide molecular-level descriptions of sodium channel function. We describe an almost barrier-less three-ion conduction mechanism involving competing knock-on and “pass-by” processes, intimately linked to signature glutamate ring protonation and structural isomerizations. These simulations have uncovered a high degree of protein flexibility, with conformational fluctuations in the pore domain involving residues central to slow-type inactivation, leading to gate collapse, helix bending, filter disruption, and changes in lipid-facing fenestrations linked to Nav drug pathways. Voltage-gated Na+ channels play an essential role in electrical signaling in the nervous system and are key pharmacological targets for a range of disorders. The recent solution of X-ray structures for the bacterial channel NavAb has provided an opportunity to study functional mechanisms at the atomic level. This channel’s selectivity filter exhibits an EEEE ring sequence, characteristic of mammalian Ca2+, not Na+, channels. This raises the fundamentally important question: just what makes a Na+ channel conduct Na+ ions? Here we explore ion permeation on multimicrosecond timescales using the purpose-built Anton supercomputer. We isolate the likely protonation states of the EEEE ring and observe a striking flexibility of the filter that demonstrates the necessity for extended simulations to study conduction in this channel. We construct free energy maps to reveal complex multi-ion conduction via knock-on and “pass-by” mechanisms, involving concerted ion and glutamate side chain movements. Simulations in mixed ionic solutions reveal relative energetics for Na+, K+, and Ca2+ within the pore that are consistent with the modest selectivity seen experimentally. We have observed conformational changes in the pore domain leading to asymmetrical collapses of the activation gate, similar to proposed inactivated structures of NavAb, with helix bending involving conserved residues that are critical for slow inactivation. These structural changes are shown to regulate access to fenestrations suggested to be pathways for lipophilic drugs and provide deeper insight into the molecular mechanisms connecting drug activity and slow inactivation.