Inward-facing conformation of a multidrug resistance MATE family transporter

Sandra Zakrzewska, Ahmad Reza Mehdipour, Viveka Nand Malviya, Tsuyoshi Nonaka, Juergen Koepke, Cornelia Muenke, Winfried Hausner, Gerhard Hummer, Schara Safarian, and Hartmut Michel
PNAS June 18, 2019 116 (25) 12275-12284; first published June 3, 2019 https://doi.org/10.1073/pnas.1904210116

  1. Contributed by Hartmut Michel, May 2, 2019 (sent for review March 12, 2019; reviewed by Raimund Dutzler and José D. Faraldo-Gómez)

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Significance

Active efflux of drugs and toxic compounds carried out by multidrug and toxic compound extrusion (MATE) family transporters is one of the strategies developed by bacterial pathogens to confer multidrug resistance. To elucidate the underlying steps of the MATE transport cycle, structures of distinct intermediates are required; however, only structures in outward-facing conformations have been available. Our approach of using native lipids allowed trapping and visualization of the MATE transporter from Pyrococcus furiosus in an inward-facing conformation. This work highlights the importance of native lipids and opens an alternative view on the function and mechanism of action for the MATE family transporters.

Abstract

Multidrug and toxic compound extrusion (MATE) transporters mediate excretion of xenobiotics and toxic metabolites, thereby conferring multidrug resistance in bacterial pathogens and cancer cells. Structural information on the alternate conformational states and knowledge of the detailed mechanism of MATE transport are of great importance for drug development. However, the structures of MATE transporters are only known in V-shaped outward-facing conformations. Here, we present the crystal structure of a MATE transporter from Pyrococcus furiosus (PfMATE) in the long-sought-after inward-facing state, which was obtained after crystallization in the presence of native lipids. Transition from the outward-facing state to the inward-facing state involves rigid body movements of transmembrane helices (TMs) 2–6 and 8–12 to form an inverted V, facilitated by a loose binding of TM1 and TM7 to their respective bundles and their conformational flexibility. The inward-facing structure of PfMATE in combination with the outward-facing one supports an alternating access mechanism for the MATE family transporters.

Footnotes

  • ↵1To whom correspondence may be addressed. Email: schara.safarian{at}biophys.mpg.de or hartmut.michel{at}biophys.mpg.de.

  • Author contributions: S.Z. prepared samples, and designed and performed research; S.Z. and S.S. implemented X-ray data acquisition and solved the 6GWH and 6FHZ structures; S.Z. performed structure refinement; A.R.M. performed MD simulations; T.N. solved the original 6HFB and 4MLB structures; S.Z. and J.K. refined the 6HFB and 4MLB structures; C.M. performed cloning, optimization of gene expression, purification, and crystallization leading to the 6HFB and 4MLB structure determination; V.N.M. performed the functional characterization of PfMATE; W.H. assisted with the cultivation of P. furiosus; S.Z., A.R.M., and S.S. prepared figures; A.R.M. prepared videos; S.Z., A.R.M., and S.S. wrote the manuscript with contributions from G.H. and H.M.; H.M. suggested experiments; G.H. and H.M. supervised the project.

  • Reviewers: R.D., University of Zurich; and J.D.F.-G., National Institutes of Health.

  • The authors declare no conflict of interest.

  • Data deposition: The structure factors and coordinates for the inward- and outward-facing states obtained by the lipidic cubic phase method were deposited in the Protein Data Bank (PDB), www.pdb.org (PDB ID codes 6FHZ and 6GWH, respectively). The crystallographic data corresponding to the outward-facing structures obtained by the vapor diffusion method were deposited in the PDB (PDB ID codes 6HFB and 4MLB).

  • This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1904210116/-/DCSupplemental.

Published under the PNAS license.

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Inward-facing conformation of a multidrug resistance MATE family transporter
Sandra Zakrzewska, Ahmad Reza Mehdipour, Viveka Nand Malviya, Tsuyoshi Nonaka, Juergen Koepke, Cornelia Muenke, Winfried Hausner, Gerhard Hummer, Schara Safarian, Hartmut Michel
Proceedings of the National Academy of Sciences Jun 2019, 116 (25) 12275-12284; DOI: 10.1073/pnas.1904210116
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