Role of the C-terminal domain in the structure and function of tetrameric sodium channels.

Claire Bagnéris,Paul G Decaen,B A Wallace,Claire E Naylor,David E Clapham,Benjamin A Hall,Christopher W M Kay

doi:10.1038/ncomms3465

Abstract

Voltage-gated sodium channels have essential roles in electrical signalling. Prokaryotic sodium channels are tetramers consisting of transmembrane (TM) voltage-sensing and pore domains, and a cytoplasmic carboxy-terminal domain. Previous crystal structures of bacterial sodium channels revealed the nature of their TM domains but not their C-terminal domains (CTDs). Here, using electron paramagnetic resonance (EPR) spectroscopy combined with molecular dynamics, we show that the CTD of the NavMs channel from Magnetococcus marinus includes a flexible region linking the TM domains to a four-helix coiled-coil bundle. A 2.9 Å resolution crystal structure of the NavMs pore indicates the position of the CTD, which is consistent with the EPR-derived structure. Functional analyses demonstrate that the coiled-coil domain couples inactivation with channel opening, and is enabled by negatively charged residues in the linker region. A mechanism for gating is proposed based on the structure, whereby splaying of the bottom of the pore is possible without requiring unravelling of the coiled-coil.

Highlights

Voltage-gated sodium channels have essential roles in electrical signalling
The crystal structure of a chimeric construct consisting of part of the distal end of the C-terminal domains (CTDs) of another NaChBac orthologue, NavSulP, attached to the end of an existing coiled-coil region of an unrelated NaK channel[14] (Fig. 1a), showed it could form a coiled-coil in that context, but it was missing the important proximal linker region between the coiled-coil and TM domain
We examine the CTD as part of the NavMs bacterial sodium channel pore[9], using continuous wave electron paramagnetic resonance and pulsed double electron– electron resonance (DEER) spectroscopy to define the supersecondary/quaternary structure of the domain

Summary

Introduction

Voltage-gated sodium channels have essential roles in electrical signalling. Prokaryotic sodium channels are tetramers consisting of transmembrane (TM) voltage-sensing and pore domains, and a cytoplasmic carboxy-terminal domain. Sequence analyses[5,6] suggested that the distal helical region would form a coiled-coil, based on the presence of the characteristic heptad repeat; the same sequence motif has been found in all prokaryotic sodium channel sequences examined to date, as well as in many related eukaryotic ion channels such as CatSper[11], TRPV1 The crystal structure of a chimeric construct consisting of part of the distal end of the CTD of another NaChBac orthologue, NavSulP, attached to the end of an existing coiled-coil region of an unrelated NaK channel[14] (Fig. 1a), showed it could form a coiled-coil in that context, but it was missing the important proximal linker region between the coiled-coil and TM domain.

Methods

Results

Conclusion