title: The Lipid Bilayer Modulates the Structure and Function of an ATP-binding Cassette Exporter.
abstract: ATP-binding cassette exporters use the energy of ATP hydrolysis to transport substrates across membranes by switching between inward- and outward-facing conformations. Essentially all structural studies of these proteins have been performed with the proteins in detergent micelles, locked in specific conformations and/or at low temperature. Here, we used luminescence resonance energy transfer spectroscopy to study the prototypical ATP-binding cassette exporter MsbA reconstituted in nanodiscs at 37 °C while it performs ATP hydrolysis. We found major differences when comparing MsbA in these native-like conditions with double electron-electron resonance data and the crystal structure of MsbA in the open inward-facing conformation. The most striking differences include a significantly smaller separation between the nucleotide-binding domains and a larger fraction of molecules with associated nucleotide-binding domains in the nucleotide-free apo state. These studies stress the importance of studying membrane proteins in an environment that approaches physiological conditions.
authors.affiliation: From the Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, Texas 79430-6551.
authors.forename: Maria E
authors.initials: ME
authors.lastname: Zoghbi
authors.affiliation: From the Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, Texas 79430-6551.
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Cooper
authors.affiliation: From the Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, Texas 79430-6551 g.altenberg@ttuhsc.edu.
authors.forename: Guillermo A
authors.initials: GA
authors.lastname: Altenberg
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D060807:Bioluminescence Resonance Energy Transfer Techniques; D004795:Enzyme Stability; D006868:Hydrolysis; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D021962:Membrane Microdomains; D008565:Membrane Proteins / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D049329:Nanostructures / Q000737:chemistry; D011487:Protein Conformation; D058767:Protein Unfolding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012486:Salmonella typhimurium / Q000201:enzymology*
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D008051:Lipid Bilayers; D008565:Membrane Proteins; C079726:MsbA protein, Bacteria; D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate
keywords: ABC transporter; FRET; LRET; MsbA; fluorescence resonance energy transfer (FRET); luminescence resonance energy transfer; membrane bilayer; multidrug transporter; nanodisc; spectroscopy