Single-molecule dynamics of gating in a neurotransmitter transporter homologue
Open Access
- 13 May 2010
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature
- Vol. 465 (7295), 188-193
- https://doi.org/10.1038/nature09057
Abstract
Neurotransmitter:Na+ symporters (NSS) remove neurotransmitters from the synapse in a reuptake process that is driven by the Na+ gradient. Drugs that interfere with this reuptake mechanism, such as cocaine and antidepressants, profoundly influence behaviour and mood. To probe the nature of the conformational changes that are associated with substrate binding and transport, we have developed a single-molecule fluorescence imaging assay and combined it with functional and computational studies of the prokaryotic NSS homologue LeuT. Here we show molecular details of the modulation of intracellular gating of LeuT by substrates and inhibitors, as well as by mutations that alter binding, transport or both. Our direct observations of single-molecule transitions, reflecting structural dynamics of the intracellular region of the transporter that might be masked by ensemble averaging or suppressed under crystallographic conditions, are interpreted in the context of an allosteric mechanism that couples ion and substrate binding to transport.Keywords
This publication has 53 references indexed in Scilit:
- Antidepressant specificity of serotonin transporter suggested by three LeuT–SSRI structuresNature Structural & Molecular Biology, 2009
- Mitigating Unwanted Photophysical Processes for Improved Single-Molecule Fluorescence ImagingBiophysical Journal, 2009
- Single-molecule observations of ribosome functionCurrent Opinion in Structural Biology, 2009
- The Mechanism of a Neurotransmitter:Sodium Symporter—Inward Release of Na+ and Substrate Is Triggered by Substrate in a Second Binding SiteMolecular Cell, 2008
- The Cys154→Gly Mutation in LacY Causes Constitutive Opening of the Hydrophilic Periplasmic PathwayJournal of Molecular Biology, 2008
- Following translation by single ribosomes one codon at a timeNature, 2008
- Protein folding studied by single-molecule FRETCurrent Opinion in Structural Biology, 2008
- Single-molecule FRET reveals sugar-induced conformational dynamics in LacYProceedings of the National Academy of Sciences of the United States of America, 2007
- Monitoring the function of membrane transport proteins in detergent-solubilized formProceedings of the National Academy of Sciences of the United States of America, 2007
- Identification of Two Distinct Hybrid State Intermediates on the RibosomeMolecular Cell, 2007