Physicochemical factors controlling the activity and energy coupling of an ionic strength-gated ATP-binding cassette (ABC) transporter

J Biol Chem. 2013 Oct 11;288(41):29862-71. doi: 10.1074/jbc.M113.499327. Epub 2013 Aug 26.

Abstract

Cells control their volume through the accumulation of compatible solutes. The bacterial ATP-binding cassette transporter OpuA couples compatible solute uptake to ATP hydrolysis. Here, we study the gating mechanism and energy coupling of OpuA reconstituted in lipid nanodiscs. We show that anionic lipids are essential both for the gating and the energy coupling. The tight coupling between substrate binding on extracellular domains and ATP hydrolysis by cytoplasmic nucleotide-binding domains allows the study of transmembrane signaling in nanodiscs. From the tight coupling between processes at opposite sides of the membrane, we infer that the ATPase activity of OpuA in nanodiscs reflects solute translocation. Intriguingly, the substrate-dependent, ionic strength-gated ATPase activity of OpuA in nanodiscs is at least an order of magnitude higher than in lipid vesicles (i.e. with identical membrane lipid composition, ionic strength, and nucleotide and substrate concentrations). Even with the chemical components the same, the lateral pressure (profile) of the nanodiscs will differ from that of the vesicles. We thus propose that membrane tension limits translocation in vesicular systems. Increased macromolecular crowding does not activate OpuA but acts synergistically with ionic strength, presumably by favoring gating interactions of like-charged surfaces via excluded volume effects.

Keywords: ABC Transporter; BC Transporter; Cell Volume Regulation; Energetics; Gating; Gating Mechanism; Ionic Strength; Macromolecular Crowding; Membrane; Membrane Reconstitution; Membrane Transport.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP-Binding Cassette Transporters / chemistry
  • ATP-Binding Cassette Transporters / genetics
  • ATP-Binding Cassette Transporters / metabolism*
  • Adenosine Triphosphatases / chemistry
  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Adenosine Triphosphate / metabolism*
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Biological Transport
  • Chemical Phenomena
  • Chromatography, Liquid
  • Energy Transfer
  • Hydrolysis
  • Lipid Bilayers / chemistry
  • Lipid Bilayers / metabolism
  • Liposomes / chemistry
  • Liposomes / metabolism
  • Mass Spectrometry
  • Membrane Lipids / chemistry
  • Membrane Lipids / metabolism
  • Nanostructures / chemistry
  • Osmolar Concentration
  • Proteolipids / chemistry
  • Proteolipids / metabolism
  • Substrate Specificity

Substances

  • ATP-Binding Cassette Transporters
  • Bacterial Proteins
  • Lipid Bilayers
  • Liposomes
  • Membrane Lipids
  • Proteolipids
  • proteoliposomes
  • Adenosine Triphosphate
  • Adenosine Triphosphatases
  • OpuA protein, Lactococcus lactis