title: Conformational change induced by ATP binding in the multidrug ATP-binding cassette transporter BmrA.
abstract: ATP-binding cassette (ABC) transporters are involved in the transport of a wide variety of substrates, and ATP-driven dimerization of their nucleotide binding domains (NBDs) has been suggested to be one of the most energetic steps of their catalytic cycle. Taking advantage of the propensity of BmrA, a bacterial multidrug resistance ABC transporter, to form stable, highly ordered ring-shaped structures [Chami et al. (2002) J. Mol. Biol. 315, 1075-1085], we show here that addition of ATP in the presence of Mg2+ prevented ring formation or destroyed the previously formed rings. To pinpoint the catalytic step responsible for such an effect, two classes of hydrolysis-deficient mutants were further studied. In contrast to hydrolytically inactive glutamate mutants that behaved essentially as the wild-type, lysine Walker A mutants formed ring-shaped structures even in the presence of ATP-Mg. Although the latter mutants still bound ATP-Mg, and even slowly hydrolyzed it for the K380R mutant, they were most likely unable to undergo a proper NBD dimerization upon ATP-Mg addition. The ATP-driven dimerization step, which was still permitted in glutamate mutants and led to a stable conformation suitable to monitor the growth of 2D crystals, appeared therefore responsible for destabilization of the BmrA ring structures. Our results provide direct visual evidence that the ATP-induced NBD dimerization triggers a conformational change large enough in BmrA to destabilize the rings, which is consistent with the assumption that this step might constitute the "power stroke" for ABC transporters.
authors.affiliation: Institut de Biologie et Chimie des Protéines, UMR 5086 CNRS-Université de Lyon 1 and IFR 128 BioSciences Gerland-Lyon Sud, 7 passage du Vercors, 69367 Lyon, France.
authors.forename: Cédric
authors.initials: C
authors.lastname: Orelle
authors.forename: Francesca
authors.initials: F
authors.lastname: Gubellini
authors.forename: Anne
authors.initials: A
authors.lastname: Durand
authors.forename: Sergio
authors.initials: S
authors.lastname: Marco
authors.forename: Daniel
authors.initials: D
authors.lastname: Lévy
authors.forename: Philippe
authors.initials: P
authors.lastname: Gros
authors.forename: Attilio
authors.initials: A
authors.lastname: Di Pietro
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Jault
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D014867:Water / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D050505:Mutant Proteins; D014867:Water; C075041:Bmr protein, Bacillus subtilis; D000255:Adenosine Triphosphate