title: Conformational Dynamics on the Extracellular Side of LeuT Controlled by Na+ and K+ Ions and the Protonation State of Glu290.
abstract: Ions play key mechanistic roles in the gating dynamics of neurotransmitter:sodium symporters (NSSs). In recent microsecond scale molecular dynamics simulations of a complete model of the dopamine transporter, a NSS protein, we observed a partitioning of K(+) ions from the intracellular side toward the unoccupied Na2 site of dopamine transporter following the release of the Na2-bound Na(+) Here we evaluate with computational simulations and experimental measurements of ion affinities under corresponding conditions, the consequences of K(+) binding in the Na2 site of LeuT, a bacterial homolog of NSS, when both Na(+) ions and substrate have left, and the transporter prepares for a new cycle. We compare the results with the consequences of binding Na(+) in the same apo system. Analysis of >50-μs atomistic molecular dynamics and enhanced sampling trajectories of constructs with Glu(290), either charged or neutral, point to the Glu(290) protonation state as a main determinant in the structural reconfiguration of the extracellular vestibule of LeuT in which a "water gate" opens through coordinated motions of residues Leu(25), Tyr(108), and Phe(253) The resulting water channel enables the binding/dissociation of the Na(+) and K(+) ions that are prevalent, respectively, in the extracellular and intracellular environments.
authors.affiliation: From the Department of Physiology and Biophysics, Weill Cornell Medical College of Cornell University, New York, New York 10065, gek2009@med.cornell.edu.
authors.forename: George
authors.initials: G
authors.lastname: Khelashvili
authors.affiliation: the Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen N, Denmark.
authors.forename: Solveig Gaarde
authors.initials: SG
authors.lastname: Schmidt
authors.affiliation: From the Department of Physiology and Biophysics, Weill Cornell Medical College of Cornell University, New York, New York 10065, the Computational Chemistry and Molecular Biophysics Unit, National Institute on Drug Abuse-Intramural Research Program, National Institutes of Health, Baltimore, Maryland 21224.
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
authors.affiliation: the Departments of Psychiatry and Pharmacology, Columbia University College of Physicians and Surgeons, New York, New York 10032, Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York 10032, and.
authors.forename: Jonathan A
authors.initials: JA
authors.lastname: Javitch
authors.affiliation: the Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen N, Denmark.
authors.forename: Ulrik
authors.initials: U
authors.lastname: Gether
authors.affiliation: the Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen N, Denmark.
authors.forename: Claus J
authors.initials: CJ
authors.lastname: Loland
authors.affiliation: From the Department of Physiology and Biophysics, Weill Cornell Medical College of Cornell University, New York, New York 10065, the Institute for Computational Biomedicine, Weill Medical College of Cornell University, New York, New York 10065.
authors.forename: Harel
authors.initials: H
authors.lastname: Weinstein
mesh_terms: D000818:Animals; D001419:Bacteria / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002414:Cations, Monovalent / Q000737:chemistry / Q000378:metabolism; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster; D018698:Glutamic Acid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D050479:Neurotransmitter Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011188:Potassium / Q000737:chemistry* / Q000378:metabolism; D017433:Protein Structure, Secondary; D012964:Sodium / Q000737:chemistry* / Q000378:metabolism; D040681:Structural Homology, Protein
chemical_list: D001426:Bacterial Proteins; D002414:Cations, Monovalent; D029721:Drosophila Proteins; D050479:Neurotransmitter Transport Proteins; D018698:Glutamic Acid; D012964:Sodium; D011188:Potassium
keywords: conformational change; membrane protein; molecular dynamics; monoamine transporter; neurotransmitter transport