title: Transmembrane gate movements in the type II ATP-binding cassette (ABC) importer BtuCD-F during nucleotide cycle.
abstract: ATP-binding cassette (ABC) transporters are ubiquitous integral membrane proteins that translocate substrates across cell membranes. The alternating access of their transmembrane domains to opposite sides of the membrane powered by the closure and reopening of the nucleotide binding domains is proposed to drive the translocation events. Despite clear structural similarities, evidence for considerable mechanistic diversity starts to accumulate within the importers subfamily. We present here a detailed study of the gating mechanism of a type II ABC importer, the BtuCD-F vitamin B(12) importer from Escherichia coli, elucidated by EPR spectroscopy. Distance changes at key positions in the translocation gates in the nucleotide-free, ATP- and ADP-bound conformations of the transporter were measured in detergent micelles and liposomes. The translocation gates of the BtuCD-F complex undergo conformational changes in line with a "two-state" alternating access model. We provide the first direct evidence that binding of ATP drives the gates to an inward-facing conformation, in contrast to type I importers specific for maltose, molybdate, or methionine. Following ATP hydrolysis, the translocation gates restore to an apo-like conformation. In the presence of ATP, an excess of vitamin B(12) promotes the reopening of the gates toward the periplasm and the dislodgment of BtuF from the transporter. The EPR data allow a productive translocation cycle of the vitamin B(12) transporter to be modeled.
authors.affiliation: Laboratory of Physical Chemistry, ETH Zurich, Wolfgang-Pauli-Strasse 10, 8093 Zurich, Switzerland.
authors.forename: Benesh
authors.initials: B
authors.lastname: Joseph
authors.forename: Gunnar
authors.initials: G
authors.lastname: Jeschke
authors.forename: Birke A
authors.initials: BA
authors.lastname: Goetz
authors.forename: Kaspar P
authors.initials: KP
authors.lastname: Locher
authors.forename: Enrica
authors.initials: E
authors.lastname: Bordignon
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D003545:Cysteine; D003593:Cytoplasm / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D004123:Dimethylamines / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism; D008958:Models, Molecular; D009068:Movement; D033902:Periplasmic Binding Proteins / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D013113:Spin Labels; D014805:Vitamin B 12 / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; C050021:BtuC peptide, E coli; C050022:BtuD peptide, E coli; C050023:BtuE protein, E coli; D003902:Detergents; D004123:Dimethylamines; D029968:Escherichia coli Proteins; D008081:Liposomes; D033902:Periplasmic Binding Proteins; D013113:Spin Labels; C014518:dodecyldimethylamine oxide; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D003545:Cysteine; D014805:Vitamin B 12