ATP binding and hydrolysis disrupt the high-affinity interaction between the heme ABC transporter HmuUV and its cognate substrate-binding protein

J Biol Chem. 2017 Sep 1;292(35):14617-14624. doi: 10.1074/jbc.M117.779975. Epub 2017 Jul 14.


Using the energy of ATP hydrolysis, ABC transporters catalyze the trans-membrane transport of molecules. In bacteria, these transporters partner with a high-affinity substrate-binding protein (SBP) to import essential micronutrients. ATP binding by Type I ABC transporters (importers of amino acids, sugars, peptides, and small ions) stabilizes the interaction between the transporter and the SBP, thus allowing transfer of the substrate from the latter to the former. In Type II ABC transporters (importers of trace elements, e.g. vitamin B12, heme, and iron-siderophores) the role of ATP remains debatable. Here we studied the interaction between the Yersinia pestis ABC heme importer (HmuUV) and its partner substrate-binding protein (HmuT). Using real-time surface plasmon resonance experiments and interaction studies in membrane vesicles, we find that in the absence of ATP the transporter and the SBP tightly bind. Substrate in excess inhibits this interaction, and ATP binding by the transporter completely abolishes it. To release the stable docked SBP from the transporter hydrolysis of ATP is required. Based on these results we propose a mechanism for heme acquisition by HmuUV-T where the substrate-loaded SBP docks to the nucleotide-free outward-facing conformation of the transporter. ATP binding leads to formation of an occluded state with the substrate trapped in the trans-membrane translocation cavity. Subsequent ATP hydrolysis leads to substrate delivery to the cytoplasm, release of the SBP, and resetting of the system. We propose that other Type II ABC transporters likely share the fundamentals of this mechanism.

Keywords: ABC transporter; ATP; ATPase; membrane protein; protein-protein interaction; surface plasmon resonance (SPR).

Publication types

  • Comparative Study

MeSH terms

  • ATP-Binding Cassette Transporters / chemistry
  • ATP-Binding Cassette Transporters / genetics
  • ATP-Binding Cassette Transporters / metabolism*
  • Adenosine Triphosphate / chemistry
  • Adenosine Triphosphate / metabolism*
  • Apoenzymes / chemistry
  • Apoenzymes / genetics
  • Apoenzymes / metabolism
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Carrier Proteins / chemistry
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism*
  • Cell Membrane / chemistry
  • Cell Membrane / metabolism
  • Dimerization
  • Heme / chemistry
  • Heme / metabolism*
  • Heme-Binding Proteins
  • Hemeproteins / chemistry
  • Hemeproteins / genetics
  • Hemeproteins / metabolism*
  • Holoenzymes / chemistry
  • Holoenzymes / genetics
  • Holoenzymes / metabolism
  • Hydrolysis
  • Immobilized Proteins / chemistry
  • Immobilized Proteins / genetics
  • Immobilized Proteins / metabolism
  • Kinetics
  • Models, Molecular*
  • Molecular Docking Simulation
  • Protein Interaction Domains and Motifs
  • Protein Multimerization
  • Receptors, Cell Surface / chemistry
  • Receptors, Cell Surface / genetics
  • Receptors, Cell Surface / metabolism
  • Recombinant Proteins
  • Surface Plasmon Resonance
  • Yersinia pestis / metabolism*


  • ATP-Binding Cassette Transporters
  • Apoenzymes
  • Bacterial Proteins
  • Carrier Proteins
  • Heme-Binding Proteins
  • Hemeproteins
  • Holoenzymes
  • Immobilized Proteins
  • Receptors, Cell Surface
  • Recombinant Proteins
  • Heme
  • Adenosine Triphosphate