Identification of a second substrate-binding site in solute-sodium symporters.

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  • Additional Information
    • Source:
      Publisher: Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology Country of Publication: United States NLM ID: 2985121R Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1083-351X (Electronic) Linking ISSN: 00219258 NLM ISO Abbreviation: J Biol Chem Subsets: MEDLINE
    • Publication Information:
      Publication: 2021- : [New York, NY] : Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology
      Original Publication: Baltimore, MD : American Society for Biochemistry and Molecular Biology
    • Subject Terms:
      Amino Acid Transport Systems, Neutral/*chemistry ; Escherichia coli Proteins/*chemistry ; Galactose/*chemistry ; Plasma Membrane Neurotransmitter Transport Proteins/*chemistry ; Sodium/*chemistry ; Sodium-Glucose Transport Proteins/*chemistry ; Symporters/*chemistry; Amino Acid Sequence ; Amino Acid Transport Systems, Neutral/metabolism ; Binding Sites ; Biological Transport ; Escherichia coli/chemistry ; Escherichia coli/metabolism ; Escherichia coli Proteins/metabolism ; Galactose/metabolism ; Kinetics ; Molecular Docking Simulation ; Molecular Dynamics Simulation ; Molecular Sequence Data ; Plasma Membrane Neurotransmitter Transport Proteins/metabolism ; Protein Binding ; Protein Folding ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Recombinant Proteins/chemistry ; Recombinant Proteins/metabolism ; Sequence Alignment ; Sodium/metabolism ; Sodium-Glucose Transport Proteins/metabolism ; Structural Homology, Protein ; Substrate Specificity ; Symporters/metabolism ; Thermodynamics ; Vibrio parahaemolyticus/chemistry ; Vibrio parahaemolyticus/metabolism
    • Abstract:
      The structure of the sodium/galactose transporter (vSGLT), a solute-sodium symporter (SSS) from Vibrio parahaemolyticus, shares a common structural fold with LeuT of the neurotransmitter-sodium symporter family. Structural alignments between LeuT and vSGLT reveal that the crystallographically identified galactose-binding site in vSGLT is located in a more extracellular location relative to the central substrate-binding site (S1) in LeuT. Our computational analyses suggest the existence of an additional galactose-binding site in vSGLT that aligns to the S1 site of LeuT. Radiolabeled galactose saturation binding experiments indicate that, like LeuT, vSGLT can simultaneously bind two substrate molecules under equilibrium conditions. Mutating key residues in the individual substrate-binding sites reduced the molar substrate-to-protein binding stoichiometry to ~1. In addition, the related and more experimentally tractable SSS member PutP (the Na(+)/proline transporter) also exhibits a binding stoichiometry of 2. Targeting residues in the proposed sites with mutations results in the reduction of the binding stoichiometry and is accompanied by severely impaired translocation of proline. Our data suggest that substrate transport by SSS members requires both substrate-binding sites, thereby implying that SSSs and neurotransmitter-sodium symporters share common mechanistic elements in substrate transport.
      (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)
    • Comments:
      Erratum in: J Biol Chem. 2015 Mar 20;290(12):7361. (PMID: 25795729)
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    • Grant Information:
      DA023694 United States DA NIDA NIH HHS; DA017293 United States DA NIDA NIH HHS; R01 DA017293 United States DA NIDA NIH HHS; R00 DA023694 United States DA NIDA NIH HHS; GM078844 United States GM NIGMS NIH HHS; R01 GM078844 United States GM NIGMS NIH HHS; K99 DA023694 United States DA NIDA NIH HHS
    • Contributed Indexing:
      Keywords: Amino Acid Transport; Computational Biology; Equilibrium Binding; Galactose; Membrane Transport; Molecular Dynamics; Scintillation Proximity Assay; Solute-Sodium Symporter; Structural Alignment; Transporter
    • Molecular Sequence:
      PDB 2A65; 3DH4; 3TT3
    • Accession Number:
      0 (Amino Acid Transport Systems, Neutral)
      0 (Escherichia coli Proteins)
      0 (Plasma Membrane Neurotransmitter Transport Proteins)
      0 (Recombinant Proteins)
      0 (Sodium-Glucose Transport Proteins)
      0 (Symporters)
      111565-48-1 (PutP protein, E coli)
      9NEZ333N27 (Sodium)
      X2RN3Q8DNE (Galactose)
    • Publication Date:
      Date Created: 20141116 Date Completed: 20150407 Latest Revision: 20210205
    • Publication Date:
      20240829
    • Accession Number:
      PMC4281715
    • Accession Number:
      10.1074/jbc.M114.584383
    • Accession Number:
      25398883