title: Glu-311 in External Loop 4 of the Sodium/Proline Transporter PutP Is Crucial for External Gate Closure.
abstract: The available structural information on LeuT and structurally related transporters suggests that external loop 4 (eL4) and the outer end of transmembrane domain (TM) 10' participate in the reversible occlusion of the outer pathway to the solute binding sites. Here, the functional significance of eL4 and the outer region of TM10' are explored using the sodium/proline symporter PutP as a model. Glu-311 at the tip of eL4, and various amino acids around the outer end of TM10' are identified as particularly crucial for function. Substitutions at these sites inhibit the transport cycle, and affect in part ligand binding. In addition, changes at selected sites induce a global structural alteration in the direction of an outward-open conformation. It is suggested that interactions between the tip of eL4 and the peptide backbone at the end of TM10' participate in coordinating conformational alterations underlying the alternating access mechanism of transport. Together with the structural information on LeuT-like transporters, the results further specify the idea that common design and functional principles are maintained across different transport families.
authors.affiliation: From the Department of Biology 1, Division of Microbiology, Ludwig Maximilians University Munich, Grosshaderner Strasse 2-4, 82152 Martinsried, Germany and.
authors.forename: Susanne
authors.initials: S
authors.lastname: Bracher
authors.affiliation: the ETH Zurich, Laboratory of Physical Chemistry, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland.
authors.forename: Kamila
authors.initials: K
authors.lastname: Guérin
authors.affiliation: the ETH Zurich, Laboratory of Physical Chemistry, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland.
authors.forename: Yevhen
authors.initials: Y
authors.lastname: Polyhach
authors.affiliation: the ETH Zurich, Laboratory of Physical Chemistry, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Jeschke
authors.affiliation: From the Department of Biology 1, Division of Microbiology, Ludwig Maximilians University Munich, Grosshaderner Strasse 2-4, 82152 Martinsried, Germany and.
authors.forename: Sophie
authors.initials: S
authors.lastname: Dittmer
authors.affiliation: From the Department of Biology 1, Division of Microbiology, Ludwig Maximilians University Munich, Grosshaderner Strasse 2-4, 82152 Martinsried, Germany and.
authors.forename: Sabine
authors.initials: S
authors.lastname: Frey
authors.affiliation: From the Department of Biology 1, Division of Microbiology, Ludwig Maximilians University Munich, Grosshaderner Strasse 2-4, 82152 Martinsried, Germany and.
authors.forename: Maret
authors.initials: M
authors.lastname: Böhm
authors.affiliation: From the Department of Biology 1, Division of Microbiology, Ludwig Maximilians University Munich, Grosshaderner Strasse 2-4, 82152 Martinsried, Germany and hjung@lmu.de.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Jung
mesh_terms: D000595:Amino Acid Sequence; D026921:Amino Acid Transport Systems, Neutral / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D005973:Glutamine / Q000737:chemistry; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D027981:Symporters / Q000737:chemistry* / Q000378:metabolism
chemical_list: D026921:Amino Acid Transport Systems, Neutral; D029968:Escherichia coli Proteins; D027981:Symporters; D005973:Glutamine; C060609:PutP protein, E coli
keywords: PutP; amino acid transport; electron paramagnetic resonance (EPR); membrane protein; proline; protein chemistry; protein conformation; secondary transport; solute/sodium symport; spin labeling