Bridge 2: Structural Dynamics of ABC Transporter

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Multidrug ATP binding cassette (ABC) exporters are ubiquitous ABC transporters that extrude cytotoxic molecules across cell membranes. Mammalian ABC transporters, such as P- glycoprotein (Pgp) and cystic fibrosis transmembrane conductance regulator (CFTR), are exclusively of the exporter class, play critical physiological roles and are associated with disease. Research will continue on the overarching goal of defining the conformational motions that transduce the ATP energy to the mechanical work of substrate translocation by ABC exporters. The design leverages robust methods and approaches, tested and refined by the Cores in phase I of the consortium, to reveal commonalities and differences in the conformational cycles of three archetypes ABC exporters selected based on specific pattern of sequence and mechanistic divergences. The specific aims seek to define the intermediate states required for ATP-powered transport, map the transition pathways and elucidate the kinetics of interconversion between states. An integrated computational and spectroscopic approach capitalizes on the expertise of five investigators, who concurrently participate in the Cores and have a track record of collaboration, to achieve the specific aims. We will use : a novel computational approach to map transition pathways, DEER spectroscopy to monitor domain movements, rapid-freeze quench to time resolve these movements and sm-FRET to detect transient intermediates and measure dwell times. Our findings will be integrated into models in order to reveal how dynamics control the function of ABC exporters.