Identification of a pre-active conformation of a pentameric channel receptor

Abstract

Pentameric ligand-gated ion channels (pLGICs) mediate fast chemical signaling through global allosteric transitions. Despite the existence of several high-resolution structures of pLGICs, their dynamical properties remain elusive. Using the proton-gated channel GLIC, we engineered multiple fluorescent reporters, each incorporating a bimane and a tryptophan/tyrosine, whose close distance causes fluorescence quenching. We show that proton application causes a global compaction of the extracellular subunit interface, coupled to an outward motion of the M2-M3 loop near the channel gate. These movements are highly similar in lipid vesicles and detergent micelles. These reorganizations are essentially completed within 2 ms and occur without channel opening at low proton concentration, indicating that they report a pre-active intermediate state in the transition pathway toward activation. This provides a template to investigate the gating of eukaryotic neurotransmitter receptors, for which intermediate states also participate in activation.DOI: http://dx.doi.org/10.7554/eLife.23955.001

The dual‐gate model for pentameric ligand‐gated ion channels activation and desensitization

“…However, recent work on the prokaryotic GLIC recently identified and characterized structurally such an intermediate. Indeed, using the tryptophan-induced fluorescence quenching method, Menny et al (2017) managed to follow the structural dynamics of a fully functional GLIC protein reconstituted into liposomes. Data show that the agonist promotes a fast quaternary compaction of the ECD in concert with a key revolving motion of the M2-M3 loop at the ECD-TMD interface.…”

Structural Basis for a Bimodal Allosteric Mechanism of General Anesthetic Modulation in Pentameric Ligand-Gated Ion Channels

“…The locally closed conformation likely represents a short-lived gating intermediate, e.g. the flip, primed, or pre-active state (Menny et al, 2017;Lev et al, 2017), that can be preferentially stabilized by chemical crosslinks or point mutations. Independent of its relevance to gating, the locally closed structure closely matches the resting state in the TMD, but it can be obtained under more flexible crystallization conditions and to higher resolution (Laurent et al, 2016); thus, it constitutes an accessible structural template for the closed pLGIC pore.…”

A tale of ligands big and small: An update on how pentameric ligand-gated ion channels interact with agonists and proteins

“…Structures of such different conformations provide key start and endpoints for in silico molecular dynamics simulations [14**]. The robustness of GLIC as a protein has allowed it to be probed in spin-labelling/EPR spectroscopy [15–18] and with fluorescence quenching [19**]. For recent reviews that cover pLGIC function in greater detail see references [20–25].…”