Functional and structural asymmetry suggest a unifying principle for catalysis in integral membrane-bound pyrophosphatases

bioRxiv (Cold Spring Harbor Laboratory)

Abstract

Abstract Membrane-bound pyrophosphatases (M-PPases) are homodimeric primary ion pumps that couple the transport of Na + - and/or H + across membranes to the hydrolysis of pyrophosphate. Their role in the virulence of protist pathogens like Plasmodium falciparum makes them an intriguing target for structural and functional studies. Here, we show the first structure of a K + -independent M-PPase, asymmetric and time-dependent substrate binding in time-resolved structures of a K + -dependent M-PPase, and demonstrate pumping-before-hydrolysis by electrometric studies. We suggest how key residues in helix 12, 13, and the exit channel loops affect ion selectivity and K + -activation due to a complex interplay of residues that are involved in subunit-subunit communication. Our findings not only explain ion selectivity in M-PPases but also why they display half-of-the-sites reactivity. Based on this we propose, for the first time, a unified and testable model for ion pumping, hydrolysis, and energy-coupling in all M-PPases, including those that pump both Na + and H + .

Authors 11

1. Jannik Strauss

   University of Leeds

   Affiliation as printed

   Astbury Centre for Structural and Molecular Biology, University of Leeds, LS2 9JT, Leeds, UK

2. Craig Wilkinson

   University of Leeds

   Affiliation as printed

   Astbury Centre for Structural and Molecular Biology, University of Leeds, LS2 9JT, Leeds, UK

3. Keni Vidilaseris

   University of Helsinki

   Affiliation as printed

   Molecular and Integrative Biosciences, Biological and Environmental Sciences, University of Helsinki, 00100 Helsinki, Finland

4. Orquídea Ribeiro

   University of Helsinki

   Affiliation as printed

   Molecular and Integrative Biosciences, Biological and Environmental Sciences, University of Helsinki, 00100 Helsinki, Finland

5. Jianing Liu

   University of Helsinki

   Affiliation as printed

   Molecular and Integrative Biosciences, Biological and Environmental Sciences, University of Helsinki, 00100 Helsinki, Finland

6. James Hillier

   University of Leeds

   Affiliation as printed

   Astbury Centre for Structural and Molecular Biology, University of Leeds, LS2 9JT, Leeds, UK

7. Anssi M. Malinen

   University of Turku

   Affiliation as printed

   Department of Life Technologies, University of Turku, FIN-20014 Turku, Finland

8. Bernadette Gehl

   University of Helsinki

   Affiliation as printed

   Molecular and Integrative Biosciences, Biological and Environmental Sciences, University of Helsinki, 00100 Helsinki, Finland

9. Lars J. C. Jeuken Aachen

   Leiden University

   Affiliation as printed

   Leiden Institute of Chemistry, University Leiden, PO Box 9502, 2300 RA Leiden, Netherlands

10. Arwen R. Pearson

    Universität Hamburg · Hamburg Centre for Ultrafast Imaging

    Affiliation as printed

    Institute for Nanostructure and Solid State Physics, Hamburg Centre for Ultrafast Imaging, Universität Hamburg, 22761 Hamburg, Germany

11. Adrian Goldman corresponding

    University of Leeds · University of Helsinki

    Affiliation as printed

    Astbury Centre for Structural and Molecular Biology, University of Leeds, LS2 9JT, Leeds, UK

    Molecular and Integrative Biosciences, Biological and Environmental Sciences, University of Helsinki, 00100 Helsinki, Finland