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Normal mode gating motions of a ligand-gated ion channel persist in a fully hydrated lipid bilayer model.

ABSTRACT: We have previously used molecular modeling and normal-mode analyses combined with experimental data to visualize a plausible model of a transmembrane ligand-gated ion channel. We also postulated how the gating motion of the channel may be affected by the presence of various ligands, especially anesthetics. As is typical for normal-mode analyses, those studies were performed in vacuo to reduce the computational complexity of the problem. While such calculations constitute an efficient way to model the large scale structural flexibility of transmembrane proteins, they can be criticized for neglecting the effects of an explicit phospholipid bilayer or hydrated environment. Here, we show the successful calculation of normal-mode motions for our model of a glycine α-1 receptor, now suspended in a fully hydrated lipid bilayer. Despite the almost uniform atomic density, the introduction of water and lipid does not grossly distort the overall gating motion. Normal-mode analysis revealed that even a fully immersed glycine α-1 receptor continues to demonstrate an iris-like channel gating motion as a low-frequency, high-amplitude natural harmonic vibration consistent with channel gating. Furthermore, the introduction of periodic boundary conditions allows the examination of simultaneous harmonic vibrations of lipid in synchrony with the protein gating motions that are compatible with reasonable lipid bilayer perturbations. While these perturbations tend to influence the overall protein motion, this work provides continued support for the iris-like motion model that characterizes gating within the family of ligand-gated ion channels.

SUBMITTER: Bertaccini EJ

PROVIDER: S-EPMC3399477 | biostudies-other | 2010 Aug

REPOSITORIES: biostudies-other

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Project description:Pentameric ligand-gated ion channels (pLGICs) are neurotransmitter-activated receptors that mediate fast synaptic transmission. In pLGICs, binding of agonist to the extracellular domain triggers a structural rearrangement that leads to the opening of an ion-conducting pore in the transmembrane domain and, in the continued presence of neurotransmitter, the channels desensitize (close). The flexible loops in each subunit that connect the extracellular binding domain (loops 2, 7, and 9) to the transmembrane channel domain (M2-M3 loop) are essential for coupling ligand binding to channel gating. Comparing the crystal structures of two bacterial pLGIC homologues, ELIC and the proton-activated GLIC, suggests channel gating is associated with rearrangements in these loops, but whether these motions accurately predict the motions in functional lipid-embedded pLGICs is unknown. Here, using site-directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy and functional GLIC channels reconstituted into liposomes, we examined if, and how far, the loops at the ECD/TMD gating interface move during proton-dependent gating transitions from the resting to desensitized state. Loop 9 moves ∼9 Å inward toward the channel lumen in response to proton-induced desensitization. Loop 9 motions were not observed when GLIC was in detergent micelles, suggesting detergent solubilization traps the protein in a nonactivatable state and lipids are required for functional gating transitions. Proton-induced desensitization immobilizes loop 2 with little change in position. Proton-induced motion of the M2-M3 loop was not observed, suggesting its conformation is nearly identical in closed and desensitized states. Our experimentally derived distance measurements of spin-labeled GLIC suggest ELIC is not a good model for the functional resting state of GLIC, and that the crystal structure of GLIC does not correspond to a desensitized state. These findings advance our understanding of the molecular mechanisms underlying pLGIC gating.

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Normal mode gating motions of a ligand-gated ion channel persist in a fully hydrated lipid bilayer model.

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