An Ion-Permeable State of the Glycine Receptor Captured by Molecular Dynamics

Cerdan, Adrien H.; Martin, Nicolas Éric; Cecchini, Marco

doi:10.1016/j.str.2018.07.019

Cited by 38 publications

(60 citation statements)

References 58 publications

(93 reference statements)

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“…Remarkably, this prediction appears to be supported by the most recent cryo-EM reconstructions of the GlyR channel isolated in complex with endogenous lipids (Yu et al, 2019). In fact and unlike D&B-open , the open channel with glycine bound features a pore radius of 2.8 Å, in agreement with previous electrophysiological recordings with polyatomic anions (Bormann et al, 1987; Rundstrom et al, 1994; Lee et al, 2003) (2.65 – 2.8 Å) and simulation results ( MD-open , 2.5 Å) (Cerdan et al, 2018), see Table 1. Moreover, in the new cryo-EM open state, the pore-lining helices M2 are tilted solely in the polar direction consistent with MD-open and in contradiction with the significant azimuthal tilting observed in the wide-open and the D&B-open structures.…”

Section: Figuresupporting

confidence: 90%

“…the wide-open structure featuring an ion pore as large as 4.4 Å in radius, and the significantly more contracted semi-open structure with a pore radius of 2.4 Å at the constriction point (Du et al, 2015). Functional annotation based on all-atom Molecular Dynamics (MD) challenged both the wide-open structure, whose pore is too wide to be blocked by picrotoxin (Gonzalez-Gutierrez et al, 2017) and is too conductive and non-selective to the size of the anion (Cerdan et al, 2018), and the semi-open structure, whose pore appeared as intermittently dehydrated in MD (Trick et al, 2016) and non-permeable to chloride at physiological conditions (Cerdan et al, 2018). Furthermore, an alternative conformation of the receptor with an intermediate pore radius (2.5 Å), which was recently captured by MD ( MD-open ), was proposed to provide a better representation of the physiologically active state (Cerdan et al, 2018).…”

Section: Figurementioning

confidence: 99%

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On the functional annotation of open-channel structures in the glycine receptor

Cerdan

Cecchini

2020

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The glycine receptor (GlyR) is by far the best-characterized pentameric ligand-gated ion channel with several high-resolution structures from X-ray crystallography, cryo-EM and modeling. Nonetheless, the significance of the currently available openpore conformations is debated due to their diversity in the pore geometry. Here, we discuss the physiological significance of existing models of the GlyR active state based on conductance and selectivity measurements by computational electrophysiology. The results support the conclusion that the original cryo-EM reconstruction of the active state obtained in detergents as well as its subsequent refinement by Molecular Dynamics simulations are likely to be nonphysiological as they feature artificially dilated ion pores. In addition, the calculations indicate that a physiologically relevant open pore configuration should be constricted within a radius of 2.5 and 2.8Å, which is consistent with previous modeling, electrophysiology measurements, and the most recent cryo-EM structures obtained in a native lipid-membrane environment.

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Section: Figuresupporting

confidence: 90%

Section: Figurementioning

confidence: 99%

On the functional annotation of open-channel structures in the glycine receptor

Cerdan

Cecchini

2020

Preprint

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“…Previous GlyR structures have shed light on the mechanism of action of full agonists and antagonists 20–23 , but structures of the receptor bound to partial agonists are still absent. Furthermore, the previously solved GlyR structures are in detergent micelles and lack the M3/M4 cytoplasmic loop, factors that likely underlie the anomalously high Po of the receptor constructs and the physiologically ‘too large’ open state of the ion channel pore 24, 25 . Here we isolated the full length GlyR with native lipids and determined high resolution structures of the receptor bound to glycine, taurine or GABA.…”

Section: Introductionmentioning

confidence: 99%

Mechanism of gating and partial agonist action in the glycine receptor

Zhu

Lape

et al. 2019

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21 22 channel states, thus explaining their lesser efficacy, yet also populate agonist-bound open and 32 desensitized states. Measurements within the neurotransmitter binding pocket, as a function of 33 bound agonist, provide a metric to correlate the extent of agonist-induced conformational changes 34 to open channel probability across the Cys-loop receptor family. 35 36 to change the conformation to a short-lived pre-open intermediate (flipped/primed), rather than 62the reduced ability to open the receptor once the intermediate is reached 12,15,16 . 63Here we investigate the structural basis of partial agonist activation in GlyR, a paradigm 64Cys-loop receptor, exploiting extensive functional characterization showing that taurine and 65 GABA act as partial agonists [17][18][19] . Previous GlyR structures have shed light on the mechanism of 66 action of full agonists and antagonists 20-23 , but structures of the receptor bound to partial agonists 67 are still absent. Furthermore, the previously solved GlyR structures are in detergent micelles and 68 lack the M3/M4 cytoplasmic loop, factors that likely underlie the anomalously high Po of the 69 receptor constructs and the physiologically 'too large' open state of the ion channel pore 24,25 . 70Here we isolated the full length GlyR with native lipids and determined high resolution structures 71 of the receptor bound to glycine, taurine or GABA. The receptors were imaged by cryo-electron 72 microscopy (cryo-EM) in MSP nanodiscs or after solubilization by styrene maleic acid (SMA) 73 copolymers 26 , showing by MD simulations that the SMA complex adopts a physiologically relevant 74 open state. Importantly, cryo-EM structures of GlyR bound to taurine or GABA reveal, for the first 75 time, agonist-bound closed states as well as open and desensitized states. 76 Glycine-bound states 77The wild-type GlyR is robustly activated by glycine and single-channel analysis of clusters 78 of receptor activity, excluding the long-lived desensitized states, reveal a maximum open 79 probability (PO) of 97% at saturating concentrations of glycine (Fig. 1a). Macroscopic current 80 recordings of GlyR activity elicited by rapid application of 10 mM glycine to outside out patches 81show that the current decays to ~ 57% of peak after 1s application because of desensitization 82 (Extended Data Fig. 1a). Taken together, single channel and macroscopic results suggest that, 83 under steady state conditions, the glycine-bound GlyR substantially populates both open and 84 desensitized states. Hence, we reconstituted detergent-purified GlyR into nanodiscs or we directly 85 page 5 of 43 extracted the receptor using SMA copolymers, followed by preparation of cryo-EM grids in the 86 presence of 10 mM glycine. 87The glycine-bound receptor in nanodiscs (GlyR-nanodisc-gly) yielded a reconstruction 88 with a single 3D class at 3.2 Å resolution (Fig. 1c, Extended Data Fig. 7, Supplementary Fig. 1, 89 Supplementary Table 1). The receptor features are well resolved and there is clear density for 90 lipi...

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“…These features are in contrast to electrophysiological measurements suggesting that the pore opening may have been exaggerated. Moreover, in MD simulations performed in the absence of backbone restraints, the open structure has been shown to rapidly collapse at the pore 19 .…”

Section: Introductionmentioning

confidence: 99%

Mechanisms of activation and desensitization of full-length glycine receptor in membranes

Kumar

Basak

Rao

et al. 2019

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Glycinergic synapses play a central role in motor control and pain processing in the central nervous system. Glycine receptors (GlyR) are key players in mediating fast inhibitory neurotransmission at these synapses. While previous high-resolution structural studies have provided insights into the molecular architecture of GlyR, several mechanistic questions pertaining to channel function are still unknown. Here, we present Cryo-EM structures of the full-length GlyR protein reconstituted into lipid nanodiscs that are captured in the unliganded (closed) and glycine-bound (open and desensitized) conformations. A comparison of the three states reveals global conformational changes underlying GlyR channel gating. The functional state assignments were validated by molecular dynamics simulations of the structures incorporated in a lipid bilayer. Observed permeation events are in agreement with the anion selectivity of the channel and the reported single-channel conductance of GlyR. These studies establish the structural basis for gating, selectivity, and single-channel conductance of GlyR in a physiological environment.

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An Ion-Permeable State of the Glycine Receptor Captured by Molecular Dynamics

Cited by 38 publications

References 58 publications

On the functional annotation of open-channel structures in the glycine receptor

On the functional annotation of open-channel structures in the glycine receptor

Mechanism of gating and partial agonist action in the glycine receptor

Mechanisms of activation and desensitization of full-length glycine receptor in membranes

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