Large domain movements through the lipid bilayer mediate substrate release and inhibition of glutamate transporters

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Abstract

Glutamate transporters are essential players in glutamatergic neurotransmission in the brain, where they maintain extracellular glutamate below cytotoxic levels and allow for rounds of transmission. The structural bases of their function are well established, particularly within a model archaeal homologue, sodium and aspartate symporter Glt_Ph. However, the mechanism of gating on the cytoplasmic side of the membrane remains ambiguous. We report Cryo-EM structures of Glt_Ph reconstituted into nanodiscs, including those structurally constrained in the cytoplasm-facing state and either apo, bound to sodium ions only, substrate, or blockers. The structures show that both substrate translocation and release involve movements of the bulky transport domain through the lipid bilayer. They further reveal a novel mode of inhibitor binding and show how solutes release is coupled to protein conformational changes. Finally, we describe how domain movements are associated with the displacement of bound lipids and significant membrane deformations, highlighting the potential regulatory role of the bilayer.

Data availability

Cryo-EM coordinate files and electron density maps have been deposited in PDB under the following codes:GltPh OFS-TBOA: PDB 6X17, EMD-21991GltPh IFS-Asp: PDB 6X15, EMD-21989GltPh IFS-TBOA: PDB 6X16, EMD-21990GltPh IFS-TFB-TBOA: PDB 6X14, EMD-21988GltPh IFS-Na: PDB 6X13, EMD-21987GltPh IFS-Apo-open: PDB 6X12, EMD-21986

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Xiaoyu Wang

Physiology and Biophysics, Weill Cornell Medicine, New York, United States

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0002-8745-8238
Olga Boudker

Department of Physiology and Biophysics, Weill Cornell Medicine, New York, United States

For correspondence
olb2003@med.cornell.edu

Competing interests
Olga Boudker, Senior editor, eLife.

"This ORCID iD identifies the author of this article:" 0000-0001-6965-0851

Funding

National Institutes of Health (R37NS085318)

Olga Boudker

National Institutes of Health (R01NS064357)

Olga Boudker

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.