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Mechanistic basis for differential inhibition of the F1Fo-ATPase by aurovertin

Authors

  • Kathryn M. Johnson,

    1. Department of Chemistry, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
    2. Graduate Program in Immunology, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
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  • Lara Swenson,

    1. Department of Chemistry, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
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  • Anthony W. Opipari Jr.,

    1. Department of Obstetrics and Gynecology, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
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  • Rolf Reuter,

    1. 3. Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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  • Nawid Zarrabi,

    1. 3. Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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  • Carol A. Fierke,

    1. Department of Chemistry, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
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  • Michael Börsch,

    1. 3. Physikalisches Institut, Universität Stuttgart, Pfaffenwaldring 57, 70569 Stuttgart, Germany
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  • Gary D. Glick

    Corresponding author
    1. Department of Chemistry, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
    2. Graduate Program in Immunology, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
    • Department of Chemistry, University of Michigan, 930 N. University Ave., Ann Arbor, MI 48109-1055
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Abstract

The mitochondrial F1Fo-ATPase performs the terminal step of oxidative phosphorylation. Small molecules that modulate this enzyme have been invaluable in helping decipher F1Fo-ATPase structure, function, and mechanism. Aurovertin is an antibiotic that binds to the β subunits in the F1 domain and inhibits F1Fo-ATPase-catalyzed ATP synthesis in preference to ATP hydrolysis. Despite extensive study and the existence of crystallographic data, the molecular basis of the differential inhibition and kinetic mechanism of inhibition of ATP synthesis by aurovertin has not been resolved. To address these questions, we conducted a series of experiments in both bovine heart mitochondria and E. coli membrane F1Fo-ATPase. Aurovertin is a mixed, noncompetitive inhibitor of both ATP hydrolysis and synthesis with lower Ki values for synthesis. At low substrate concentrations, inhibition is cooperative suggesting a stoichiometry of two aurovertin per F1Fo-ATPase. Furthermore, aurovertin does not completely inhibit the ATP hydrolytic activity at saturating concentrations. Single-molecule experiments provide evidence that the residual rate of ATP hydrolysis seen in the presence of saturating concentrations of aurovertin results from a decrease in the binding change mechanism by hindering catalytic site interactions. The results from these studies should further the understanding of how the F1Fo-ATPase catalyzes ATP synthesis and hydrolysis. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 830–840, 2009.

This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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