title: Structure of the SLC4 transporter Bor1p in an inward-facing conformation.
abstract: Bor1p is a secondary transporter in yeast that is responsible for boron transport. Bor1p belongs to the SLC4 family which controls bicarbonate exchange and pH regulation in animals as well as borate uptake in plants. The SLC4 family is more distantly related to members of the Amino acid-Polyamine-organoCation (APC) superfamily, which includes well studied transporters such as LeuT, Mhp1, AdiC, vSGLT, UraA, SLC26Dg. Their mechanism generally involves relative movements of two domains: a core domain that binds substrate and a gate domain that in many cases mediates dimerization. To shed light on conformational changes governing transport by the SLC4 family, we grew helical membrane crystals of Bor1p from Saccharomyces mikatae and determined a structure at ∼6 Å resolution using cryo-electron microscopy. To evaluate the conformation of Bor1p in these crystals, a homology model was built based on the related anion exchanger from red blood cells (AE1). This homology model was fitted to the cryo-EM density map using the Molecular Dynamics (MD) Flexible Fitting method and then relaxed by all-atom MD simulation in explicit solvent and membrane. Mapping of water accessibility indicates that the resulting structure represents an inward-facing conformation. Comparisons of the resulting Bor1p model with the X-ray structure of AE1 in an outward-facing conformation, together with MD simulations of inward-facing and outward-facing Bor1p models, suggest rigid body movements of the core domain relative to the gate domain. These movements are consistent with the rocking-bundle transport mechanism described for other members of the APC superfamily.
authors.affiliation: Skirball Institute for Biomolecular Medicine, Department of Cell Biology, New York University School of Medicine, New York, New York, 10016.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Coudray
authors.affiliation: Department of Physics, Arizona State University, Tempe, Arizona, 85287.
authors.forename: Sean
authors.initials: S
authors.lastname: L Seyler
authors.affiliation: Skirball Institute for Biomolecular Medicine, Department of Cell Biology, New York University School of Medicine, New York, New York, 10016.
authors.forename: Ralph
authors.initials: R
authors.lastname: Lasala
authors.affiliation: Skirball Institute for Biomolecular Medicine, Department of Cell Biology, New York University School of Medicine, New York, New York, 10016.
authors.forename: Zhening
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Department of Pediatrics, University of Rochester Medical Center, Rochester, New York, 14652.
authors.forename: Kathy M
authors.initials: KM
authors.lastname: Clark
authors.affiliation: Department of Pediatrics, University of Rochester Medical Center, Rochester, New York, 14652.|Department of Biochemistry and Biophysics, University of Rochester Medical Center, Rochester, New York, 14652.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Dumont
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia, 20147.
authors.forename: Alexis
authors.initials: A
authors.lastname: Rohou
authors.affiliation: Department of Physics, Arizona State University, Tempe, Arizona, 85287.|Center for Biological Physics, Arizona State University, Tempe, Arizona, 85287.
authors.forename: Oliver
authors.initials: O
authors.lastname: Beckstein
authors.affiliation: Skirball Institute for Biomolecular Medicine, Department of Cell Biology, New York University School of Medicine, New York, New York, 10016.
authors.forename: David L
authors.initials: DL
authors.lastname: Stokes
mesh_terms: D001457:Anion Exchange Protein 1, Erythrocyte / Q000648:ultrastructure; D020285:Cryoelectron Microscopy / Q000379:methods*; D005656:Fungal Proteins / Q000648:ultrastructure*; D026901:Membrane Transport Proteins / Q000648:ultrastructure*; D056004:Molecular Dynamics Simulation; D012440:Saccharomyces / Q000648:ultrastructure*; D040681:Structural Homology, Protein
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D005656:Fungal Proteins; D026901:Membrane Transport Proteins
keywords: 2D crystallization; alternating access mechanism; anion exchanger family; boron transporter; cryo-EM; helical reconstruction; homology modeling; molecular dynamics; rocking-bundle mechanism; secondary transport