Substrate transport and anion permeation proceed through distinct pathways in glutamate transporters
- School of Medicine, University of Pittsburgh, United States
- National Institutes of Health, United States
Abstract
Advances in structure-function analyses and computational biology have enabled a deeper understanding of how excitatory amino acid transporters (EAATs) mediate chloride permeation and substrate transport. However, the mechanism of structural coupling between these functions remains to be established. Using a combination of molecular modeling, substituted cysteine accessibility, electrophysiology and glutamate uptake assays, we identified a chloride-channeling conformer, iChS, transiently accessible as EAAT1 reconfigures from substrate/ion-loaded into a substrate-releasing conformer. Opening of the anion permeation path in this iChS is controlled by the elevator-like movement of the substrate-binding core, along with its wall that simultaneously lines the anion permeation path (global); and repacking of a cluster of hydrophobic residues near the extracellular vestibule (local). Moreover, our results demonstrate that stabilization of iChS by chemical modifications favors anion channeling at the expense of substrate transport, suggesting a mutually exclusive regulation mediated by the movement of the flexible wall lining the two regions.
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Funding
NIH (P41GM103712)
- Ivet Bahar
NIH (P30DA035778)
- Ivet Bahar
NIH (5R01GM099738)
- Ivet Bahar
NIH (MH002946)
- Susan G Amara
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
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Substrate transport and anion permeation proceed through distinct pathways in glutamate transporters
eLife 6:e25850.
https://doi.org/10.7554/eLife.25850
