Flexibility in the ABC transporter MsbA: alternating access with a twist. Proc Natl Acad Sci USA

Departments of Cell Biology and Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, CB-105, La Jolla, CA 92037, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 12/2007; 104(48):19005-10. DOI: 10.1073/pnas.0709388104
Source: PubMed


ATP-binding cassette (ABC) transporters are integral membrane proteins that translocate a wide variety of substrates across cellular membranes and are conserved from bacteria to humans. Here we compare four x-ray structures of the bacterial ABC lipid flippase, MsbA, trapped in different conformations, two nucleotide-bound structures and two in the absence of nucleotide. Comparison of the nucleotide-free conformations of MsbA reveals a flexible hinge formed by extracellular loops 2 and 3. This hinge allows the nucleotide-binding domains to disassociate while the ATP-binding half sites remain facing each other. The binding of the nucleotide causes a packing rearrangement of the transmembrane helices and changes the accessibility of the transporter from cytoplasmic (inward) facing to extracellular (outward) facing. The inward and outward openings are mediated by two different sets of transmembrane helix interactions. Altogether, the conformational changes between these structures suggest that large ranges of motion may be required for substrate transport.

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Available from: Christopher B Roth, May 19, 2014
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    • "Resolutions were $25–40 A ˚ . The 3D structures were compared with a 3D reference set derived from a linear interpolation of X-ray structures (Ward et al., 2007) calculated using the Chimera software package (Pettersen et al., 2004). The number of particles contributing to the individual volumes was used to determine the percentages provided. "
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    ABSTRACT: ATP-binding cassette (ABC) exporters are ubiquitously found in all kingdoms of life and their members play significant roles in mediating drug pharmacokinetics and multidrug resistance in the clinic. Significant questions and controversies remain regarding the relevance of their conformations observed in X-ray structures, their structural dynamics, and mechanism of transport. Here, we used single particle electron microscopy (EM) to delineate the entire conformational spectrum of two homologous ABC exporters (bacterial MsbA and mammalian P-glycoprotein) and the influence of nucleotide and substrate binding. Newly developed amphiphiles in complex with lipids that support high protein stability and activity enabled EM visualization of individual complexes in a membrane-mimicking environment. The data provide a comprehensive view of the conformational flexibility of these ABC exporters under various states and demonstrate not only similarities but striking differences between their mechanistic and energetic regulation of conformational changes. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Structure 01/2015; 23(3). DOI:10.1016/j.str.2014.12.013 · 5.62 Impact Factor
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    • "CH2 interacts with the Q-loop of the opposite half-transporter in trans [65]. CH1 senses the nucleotide binding status during the catalytic cycle by contacting the Q-loop of both NBDs [66] (in cis and in trans) in the outward-facing conformation, whereas it interacts with the own NBD in cis in the inward-facing conformation [35] [66] [67]. "
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    ABSTRACT: Protein homeostasis results in a steady supply of peptides, which are further degraded to fuel protein synthesis or metabolic needs of the cell. In higher vertebrates, a small fraction of the resulting peptidome, however, is translocated into the ER lumen by the transporter associated with antigen processing (TAP). Antigenic peptides are guided to major histocompatibility complex class I (MHC I) molecules and are finally displayed on the cell surface, where they mount an adaptive immune response against viral infected or malignantly transformed cells. Here, we review the structural organization and the molecular mechanism of this specialized antigen translocon. We discuss how the ATP-binding cassette (ABC) transporter TAP communicates and cooperates within the multi-component peptide-loading machinery, mediating the proper assembly and editing of kinetically stable peptide/MHC I complexes. In light of its important role within the MHC I antigen processing pathway, TAP is a prime target for viral immune evasion strategies, and we summarize how this antigen translocation machinery is sabotaged by viral factors. Finally, we compare TAP with other ABC systems that facilitate peptide translocation.
    Journal of Molecular Biology 09/2014; 427(5). DOI:10.1016/j.jmb.2014.09.006 · 4.33 Impact Factor
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    • "3B5W (5.30 Å) Inward, open (apo) None (DDM) Vibrio cholerae [54] 3B5X (5.50 Å) Inward, closed (apo) None (DDM) Salmonella typhimurium [54] "
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    ABSTRACT: Background: ABC transporters ubiquitously found in all kingdoms of life move a broad range of solutes across membranes. Crystal structures of four distinct types of ABC transport systems have been solved, shedding light on different conformational states within the transport process. Briefly, ATP-dependent flipping between inward- and outward-facing conformations allows directional transport of various solutes. Scope of review: The heterodimeric transporter associated with antigen processing TAP1/2 (ABCB2/3) is a crucial element of the adaptive immune system. The ABC transport complex shuttles proteasomal degradation products into the endoplasmic reticulum. These antigenic peptides are loaded onto major histocompatibility complex class I molecules and presented on the cell surface. We detail the functional modules of TAP, its ATPase and transport cycle, and its interaction with and modulation by other cellular components. In particular, we emphasize how viral factors inhibit TAP activity and thereby prevent detection of the infected host cell by cytotoxic T-cells. Major conclusions: Merging functional details on TAP with structural insights from related ABC transporters refines the understanding of solute transport. Although human ABC transporters are extremely diverse, they still may employ conceptually related transport mechanisms. Appropriately, we delineate a working model of the transport cycle and how viral factors arrest TAP in distinct conformations. General significance: Deciphering the transport cycle of human ABC proteins is the major issue in the field. The defined peptidic substrate, various inhibitory viral factors, and its role in adaptive immunity provide unique tools for the investigation of TAP, making it an ideal model system for ABC transporters in general. This article is part of a Special Issue entitled Structural biochemistry and biophysics of membrane proteins.
    Biochimica et Biophysica Acta (BBA) - General Subjects 06/2014; 1850(3). DOI:10.1016/j.bbagen.2014.05.022 · 4.38 Impact Factor
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