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Packing of the extracellular domain hydrophobic core has evolved to facilitate pentameric ligand-gated ion channel function.

ABSTRACT: Protein function depends on conformational flexibility and folding stability. Loose packing of hydrophobic cores is not infrequent in proteins, as the enhanced flexibility likely contributes to their biological function. Here, using experimental and computational approaches, we show that eukaryotic pentameric ligand-gated ion channels are characterized by loose packing of their extracellular domain ?-sandwich cores, and that loose packing contributes to their ability to rapidly switch from closed to open channel states in the presence of ligand. Functional analyses of GABA(A) receptors show that increasing the ?-core packing disrupted GABA-mediated currents, with impaired GABA efficacy and slowed GABA current activation and desensitization. We propose that loose packing of the hydrophobic ?-core developed as an evolutionary strategy aimed to facilitate the allosteric mechanisms of eukaryotic pentameric ligand-gated ion channels.

SUBMITTER: Dellisanti CD

PROVIDER: S-EPMC3030369 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Packing of the extracellular domain hydrophobic core has evolved to facilitate pentameric ligand-gated ion channel function.

Dellisanti Cosma D CD Hanson Sonya M SM Chen Lin L Czajkowski Cynthia C

The Journal of biological chemistry 20101122 5

Protein function depends on conformational flexibility and folding stability. Loose packing of hydrophobic cores is not infrequent in proteins, as the enhanced flexibility likely contributes to their biological function. Here, using experimental and computational approaches, we show that eukaryotic pentameric ligand-gated ion channels are characterized by loose packing of their extracellular domain β-sandwich cores, and that loose packing contributes to their ability to rapidly switch from close ...[more]

PMID: 21098036

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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.

Dataset Information

Packing of the extracellular domain hydrophobic core has evolved to facilitate pentameric ligand-gated ion channel function.

Publications

Packing of the extracellular domain hydrophobic core has evolved to facilitate pentameric ligand-gated ion channel function.

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