Structural Basis for Allosteric Coupling at the Membrane-Protein Interface in Gloeobacter violaceus Ligand-gated Ion Channel (GLIC)

Phanindra Velisetty,Sreevatsa V Chalamalasetti,Sudha Chakrapani

doi:10.1074/jbc.m113.523050

Phanindra Velisetty, Sreevatsa V Chalamalasetti + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m113.523050

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Abstract

Ligand binding at the extracellular domain of pentameric ligand-gated ion channels initiates a relay of conformational changes that culminates at the gate within the transmembrane domain. The interface between the two domains is a key structural entity that governs gating. Molecular events in signal transduction at the interface are poorly defined because of its intrinsically dynamic nature combined with functional modulation by membrane lipid and water vestibules. Here we used electron paramagnetic resonance spectroscopy to delineate protein motions underlying Gloeobacter violaceus ligand-gated ion channel gating in a membrane environment and report the interface conformation in the closed and the desensitized states. Extensive intrasubunit interactions were observed in the closed state that are weakened upon desensitization and replaced by newer intersubunit contacts. Gating involves major rearrangements of the interfacial loops, accompanied by reorganization of the protein-lipid-water interface. These structural changes may serve as targets for modulation of gating by lipids, alcohols, and amphipathic drug molecules.

Highlights

Allosteric mechanisms in ligand-gated ion-channels that couple neurotransmitter binding to channel opening are poorly understood
Ligand binding at the extracellular domain of pentameric ligand-gated ion channels initiates a relay of conformational changes that culminates at the gate within the transmembrane domain
We used electron paramagnetic resonance spectroscopy to delineate protein motions underlying Gloeobacter violaceus ligand-gated ion channel gating in a membrane environment and report the interface conformation in the closed and the desensitized states

Summary

Background

Allosteric mechanisms in ligand-gated ion-channels (pLGIC) that couple neurotransmitter binding to channel opening are poorly understood. JANUARY 31, 2014 VOLUME 289 NUMBER 5 within the extracellular domain (ECD) initiates conformational changes that traverse 50 – 60 Å across the channel and result in pore opening at the transmembrane domain (TMD) This allosteric transition is believed to involve a large-scale global change in the quaternary structure [1,2,3,4], the molecular details of protein motions remain unclear. The packing of helical bundles within the TMD supports distinct water pockets, referred to as the intrasubunit and intersubunit cavities These water vestibules form the binding site for several allosteric ligands and therapeutic agents [11, 26, 27] that modulate channel gating. A comparison of GLIC and ELIC structures shows small differences in the relative positioning of their interfacial loops (Fig. 1A) Such small changes in conformation are unexpected, given the dramatic functional impact of perturbations in this region in other members of the family We analyze these findings in the light of available gating models on the basis of x-ray structures and electron micrographs

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION