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Structural basis for the drug extrusion mechanism by a MATE multidrug transporter

Nature volume 496, pages 247–251 (2013)Cite this article

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An Author Correction to this article was published on 30 January 2020

Abstract

Multidrug and toxic compound extrusion (MATE) family transporters are conserved in the three primary domains of life (Archaea, Bacteria and Eukarya), and export xenobiotics using an electrochemical gradient of H+ or Na+ across the membrane1,2. MATE transporters confer multidrug resistance to bacterial pathogens3,4,5,6 and cancer cells7, thus causing critical reductions in the therapeutic efficacies of antibiotics and anti-cancer drugs, respectively. Therefore, the development of MATE inhibitors has long been awaited in the field of clinical medicine8,9. Here we present the crystal structures of the H+-driven MATE transporter from Pyrococcus furiosus in two distinct apo-form conformations, and in complexes with a derivative of the antibacterial drug norfloxacin and three in vitro selected thioether-macrocyclic peptides, at 2.1–3.0 Å resolutions. The structures, combined with functional analyses, show that the protonation of Asp 41 on the amino (N)-terminal lobe induces the bending of TM1, which in turn collapses the N-lobe cavity, thereby extruding the substrate drug to the extracellular space. Moreover, the macrocyclic peptides bind the central cleft in distinct manners, which correlate with their inhibitory activities. The strongest inhibitory peptide that occupies the N-lobe cavity may pave the way towards the development of efficient inhibitors against MATE transporters.

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Figure 1: Overall structures of PfMATE.
Figure 2: Mutational analyses of PfMATE.
Figure 3: Complex structure of PfMATE and drug substrate.
Figure 4: Complex structures with the macrocyclic peptides.

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Primary accessions

Protein Data Bank

Data deposits

The coordinates and structure factors for the P. furiosus apo MATE (‘straight’ and ‘bent’ conformations), P26A mutant, Br-NRF-bound MATE and peptide-bound MATEs have been deposited in the Protein Data Bank, under the accession numbers 3VVN, 3VVO, 3W4T, 3VVP, 3VVQ, 3VVR and 3VVS, respectively.

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Acknowledgements

We are grateful to the beam-line staff at BL32XU of SPring-8 for assistance in data collection (proposals 2011B1062, 2012A1087 and 2012B1161), and the RIKEN BioResource Center (Ibaraki, Japan) for providing the P. furiosus genomic DNA. We are also grateful to N. Dohmae and Y. Sugita (RIKEN Advanced Science Institute, Japan) for discussions. This work was supported by the Japan Society for the Promotion of Science (JSPS) through its ‘Funding Program for World-Leading Innovative R&D on Science and Technology (FIRST program)’ to O.N.; by the Core Research for Evolutional Science and Technology Program ‘The Creation of Basic Medical Technologies to Clarify and Control the Mechanisms Underlying Chronic Inflammation’ of Japan Science and Technology Agency to O.N.; and by a Grant-in-Aid for Scientific Research (S) (24227004) and a Grant-in-Aid for Young Scientists (A) (22687007) from MEXT to O.N. and R.I., respectively. This work was also supported by a JSPS Grant-in-Aid for the Specially Promoted Research (21000005) and MEXT Platform for Drug Discovery, Informatics, and Structural Life Science to H.S., and a Grant-in-Aid for JSPS post-doctoral fellows to C.J.H. (P11344).

Author information

Author notes

  1. Yoshiki Tanaka

    Present address: Present address: Department of Medical Chemistry and Cell Biology, Faculty of Medicine, Kyoto University, Konoe-cho, Yoshida, Sakyo-ku, Kyoto, 606-8501, Japan.,

Authors and Affiliations

  1. RIKEN Advanced Science Institute, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan,

    Yoshiki Tanaka, Hideaki E. Kato, Motoyuki Hattori, Kaoru Kumazaki, Tomoya Tsukazaki, Ryuichiro Ishitani & Osamu Nureki

  2. Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan,

    Yoshiki Tanaka, Hideaki E. Kato, Motoyuki Hattori, Kaoru Kumazaki, Tomoya Tsukazaki, Ryuichiro Ishitani & Osamu Nureki

  3. Department of Chemistry, Graduate School of Science, The University of Tokyo, 7-3-1 Bunkyo-ku, Tokyo 113-0033, Japan,

    Christopher J. Hipolito, Takashi Higuchi, Takayuki Katoh & Hiroaki Suga

  4. Department of Bioengineering Sciences, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan,

    Andrés D. Maturana

  5. Department of Medical Genome Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8562, Japan,

    Koichi Ito

  6. Department of Genome Applied Microbiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University, Okayama, 700-8530, Tsushima, Japan

    Teruo Kuroda

  7. Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan,

    Motoyuki Hattori & Tomoya Tsukazaki

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Contributions

Y.T. expressed and purified PfMATE for crystallization, collected the diffraction data, solved the structures and made the mutants for functional analyses. C.J.H. performed selections, syntheses and inhibition assays of macrocyclic peptides. A.D.M. performed the fluorescence analysis. K.I. performed growth complementation tests. T.K. performed drug susceptibility tests. T.H. synthesized Br-NRF. K.K. and H.E.K. assisted with data collection. M.H. and T.T. contributed to the early stage of the project. Y.T., C.J.H., H.E.K., R.I. and O.N. wrote the manuscript. H.S. and O.N. directed and supervised all of the research.

Corresponding authors

Correspondence to Hiroaki Suga or Osamu Nureki.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains a Supplementary Discussion, Supplementary Tables 1-2, Supplementary Figures 1-10 and Supplementary References. (PDF 5096 kb)

Proton-dependent drug extrusion by MATE

The structural transition between the “straight” and “bent” conformations, viewed from the extracellular and membrane sides. (MP4 3137 kb)

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Tanaka, Y., Hipolito, C., Maturana, A. et al. Structural basis for the drug extrusion mechanism by a MATE multidrug transporter. Nature 496, 247–251 (2013). https://doi.org/10.1038/nature12014

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