Journal of Biological Chemistry
Volume 291, Issue 8, 19 February 2016, Pages 3682-3692
Journal home page for Journal of Biological Chemistry

Molecular Biophysics
Gain-of-Function Mutation W493R in the Epithelial Sodium Channel Allosterically Reconfigures Intersubunit Coupling*

https://doi.org/10.1074/jbc.M115.678052Get rights and content
Under a Creative Commons license
open access

Sodium absorption in epithelial cells is rate-limited by the epithelial sodium channel (ENaC) activity in lung, kidney, and the distal colon. Pathophysiological conditions, such as cystic fibrosis and Liddle syndrome, result from water-electrolyte imbalance partly due to malfunction of ENaC regulation. Because the quaternary structure of ENaC is yet undetermined, the bases of pathologically linked mutations in ENaC subunits α, β, and γ are largely unknown. Here, we present a structural model of heterotetrameric ENaC α1βα2γ that is consistent with previous cross-linking results and site-directed mutagenesis experiments. By using this model, we show that the disease-causing mutation αW493R rewires structural dynamics of the intersubunit interfaces α1β and α2γ. Changes in dynamics can allosterically propagate to the channel gate. We demonstrate that cleavage of the γ-subunit, which is critical for full channel activation, does not mediate activation of ENaC by αW493R. Our molecular dynamics simulations led us to identify a channel-activating electrostatic interaction between α2Arg-493 and γGlu-348 at the α2γ interface. By neutralizing a sodium-binding acidic patch at the α1β interface, we reduced ENaC activation of αW493R by more than 2-fold. By combining homology modeling, molecular dynamics, cysteine cross-linking, and voltage clamp experiments, we propose a dynamics-driven model for the gain-of-function in ENaC by αW493R. Our integrated computational and experimental approach advances our understanding of structure, dynamics, and function of ENaC in its disease-causing state.

electrophysiology
ion channel
molecular docking
molecular dynamics
protein-protein interaction
allosteric activation
channelopathies
tetramer model

Cited by (0)

*

This work was supported by National Institutes of Health Program Project Grant P01 HL 110873-04. The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

This article was selected as a Paper of the Week.

This article contains supplemental movies.

4

M. Shobair, P. Kota, and N. V. Dokholyan, unpublished data.

1

A Howard Hughes Medical Institute International Student Research Fellow.