title: Ion/substrate-dependent conformational dynamics of a bacterial homolog of neurotransmitter:sodium symporters.
abstract: Crystallographic, computational and functional analyses of LeuT have revealed details of the molecular architecture of Na(+)-coupled transporters and the mechanistic nature of ion/substrate coupling, but the conformational changes that support a functional transport cycle have yet to be described fully. We have used site-directed spin labeling and electron paramagnetic resonance (EPR) analysis to capture the dynamics of LeuT in the region of the extracellular vestibule associated with the binding of Na(+) and leucine. The results outline the Na(+)-dependent formation of a dynamic outward-facing intermediate that exposes the primary substrate binding site and the conformational changes that occlude this binding site upon subsequent binding of the leucine substrate. Furthermore, the binding of the transport inhibitors tryptophan, clomipramine and octyl-glucoside is shown to induce structural changes that distinguish the resulting inhibited conformation from the Na(+)/leucine-bound state.
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
authors.forename: Derek P
authors.initials: DP
authors.lastname: Claxton
authors.forename: Matthias
authors.initials: M
authors.lastname: Quick
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
authors.forename: Fernanda Delmondes
authors.initials: FD
authors.lastname: de Carvalho
authors.forename: Harel
authors.initials: H
authors.lastname: Weinstein
authors.forename: Jonathan A
authors.initials: JA
authors.lastname: Javitch
authors.forename: Hassane S
authors.initials: HS
authors.lastname: McHaourab
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D007930:Leucine / Q000378:metabolism*; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D019906:Nuclear Magnetic Resonance, Biomolecular; D050480:Plasma Membrane Neurotransmitter Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D040681:Structural Homology, Protein
chemical_list: D001426:Bacterial Proteins; D050480:Plasma Membrane Neurotransmitter Transport Proteins; D012964:Sodium; D007930:Leucine