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PLoS Comput Biol. 2016 Nov 11;12(11):e1005197. doi: 10.1371/journal.pcbi.1005197. eCollection 2016 Nov.

The Environment Shapes the Inner Vestibule of LeuT.

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Medical University of Vienna, Center for Physiology and Pharmacology, Institute of Pharmacology, Vienna, Austria.
University of Copenhagen, Faculty of Health and Medical Sciences Denmark, Department of Neuroscience and Pharmacology, Copenhagen, Denmark.
University of Copenhagen, Faculty of Health Sciences Denmark, Department of Biomedical Sciences, Copenhagen, Denmark.
Ludwig Maximilians University Munich, Department of Pharmacy, Center of Drug Research, Munich, Germany.


Human neurotransmitter transporters are found in the nervous system terminating synaptic signals by rapid removal of neurotransmitter molecules from the synaptic cleft. The homologous transporter LeuT, found in Aquifex aeolicus, was crystallized in different conformations. Here, we investigated the inward-open state of LeuT. We compared LeuT in membranes and micelles using molecular dynamics simulations and lanthanide-based resonance energy transfer (LRET). Simulations of micelle-solubilized LeuT revealed a stable and widely open inward-facing conformation. However, this conformation was unstable in a membrane environment. The helix dipole and the charged amino acid of the first transmembrane helix (TM1A) partitioned out of the hydrophobic membrane core. Free energy calculations showed that movement of TM1A by 0.30 nm was driven by a free energy difference of ~15 kJ/mol. Distance measurements by LRET showed TM1A movements, consistent with the simulations, confirming a substantially different inward-open conformation in lipid bilayer from that inferred from the crystal structure.

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