Structural biology of membrane transporters involved in the efflux mechanism of organic molecules

• Project/Area Number: 17H03640
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: The University of Tokyo
• Principal Investigator: Ishitani Ryuichiro 東京大学, 大学院理学系研究科(理学部), 特任教授 (90361568)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥17,420,000 (Direct Cost: ¥13,400,000、Indirect Cost: ¥4,020,000); Fiscal Year 2019: ¥5,460,000 (Direct Cost: ¥4,200,000、Indirect Cost: ¥1,260,000); Fiscal Year 2018: ¥5,460,000 (Direct Cost: ¥4,200,000、Indirect Cost: ¥1,260,000); Fiscal Year 2017: ¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000)
• Keywords: Ｘ線結晶構造解析 / 膜タンパク質 / 多剤耐性菌 / 薬物動態 / 結晶構造解析 / 膜蛋白質 / 輸送体 / 多剤耐性因子

Outline of Final Research Achievements

Living organisms are constantly exposed to toxic organic molecules, and cells have a mechanism to excrete them as a defensive measure. The molecular entity of this efflux mechanism is the membrane protein, the "efflux transporter". In this study, we analyzed the structure of MATE, a multidrug efflux transporter, focusing on the efflux mechanism of organic molecules. For eukaryotic MATE, we determined the structure of the outward opening state of the MATE transporter AtDTX14, from Arabidopsis, and verified the transport mechanism by structure-based mutational analysis. We also analyzed the structure of VcmN, a MATE from a pathogenic bacterium. The results of this structural analysis suggested that VcmN extrudes the substrate by H+-motive force-dependent TM1 bending.

Academic Significance and Societal Importance of the Research Achievements

生体は常に有毒な有機分子にさらされているが、細胞は防御策としてこれらを排出する機構を有している。これらの排出にかかわるMATE輸送体は細胞内外の物質の分布を制御する役割を担うため、その輸送機構を解明することは生命現象を理解するうえで必須であり、さらに医薬への応用などにおいても重要である。

Research Products

• [Int'l Joint Research] Vanderbilt University Medical Center/Nashville, TN 37232(米国)
• [Journal Article] Crystal structure of Drosophila Piwi. (2020)
  • Author(s): Yamaguchi, S., Oe, A., Nishida, KM., Yamashita, K., Kajiya, A., Hirano, S., Matsumoto, N., Dohmae, N., Ishitani, R., Saito, K., Siomi, H., Nishimasu, H., Siomi, MC., and Nureki, O.
  • Journal Title: Nature Communications Volume: 11 Issue: 1 Pages: 858-858
  • DOI: 10.1038/s41467-020-14687-1
  • Peer Reviewed / Open Access
• [Journal Article] Identification of Potent In Vivo Autotaxin Inhibitors that Bind to Both Hydrophobic Pockets and Channels in the Catalytic Domain (2020)
  • Author(s): Kawaguchi Mitsuyasu、Okabe Takayoshi、Okudaira Shinichi、Hama Kotaro、Kano Kuniyuki、Nishimasu Hiroshi、Nakagawa Hidehiko、Ishitani Ryuichiro、Kojima Hirotatsu、Nureki Osamu、Aoki Junken、Nagano Tetsuo
  • Journal Title: Journal of Medicinal Chemistry Volume: 63 Issue: 6 Pages: 3188-3204
  • DOI: 10.1021/acs.jmedchem.9b01967
  • Peer Reviewed / Open Access
• [Journal Article] Cryo-EM structure of the volume-regulated anion channel LRRC8D isoform identifies features important for substrate permeation (2020)
  • Author(s): Nakamura Ryoki、Numata Tomohiro、Kasuya Go、Yokoyama Takeshi、Nishizawa Tomohiro、Kusakizako Tsukasa、Kato Takafumi、Hagino Tatsuya、Dohmae Naoshi、Inoue Masato、Watanabe Kengo、Ichijo Hidenori、Kikkawa Masahide、Shirouzu Mikako、Jentsch Thomas J.、Ishitani Ryuichiro、Okada Yasunobu、Nureki Osamu
  • Journal Title: Communications Biology Volume: 3 Issue: 1 Pages: 240-240
  • DOI: 10.1038/s42003-020-0951-z
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Cryo-EM structure of the human L-type amino acid transporter 1 in complex with glycoprotein CD98hc (2019)
  • Author(s): Lee Yongchan、Wiriyasermkul Pattama、Jin Chunhuan、Quan Lili、Ohgaki Ryuichi、Okuda Suguru、Kusakizako Tsukasa、Nishizawa Tomohiro、Oda Kazumasa、Ishitani Ryuichiro、Yokoyama Takeshi、Nakane Takanori、Shirouzu Mikako、Endou Hitoshi、Nagamori Shushi、Kanai Yoshikatsu、Nureki Osamu
  • Journal Title: Nature Structural & Molecular Biology Volume: 26 Issue: 6 Pages: 510-517
  • DOI: 10.1038/s41594-019-0237-7
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural biology of the multidrug and toxic compound extrusion superfamily transporters (2019)
  • Author(s): Kusakizako Tsukasa、Miyauchi Hirotake、Ishitani Ryuichiro、Nureki Osamu
  • Journal Title: Biochimica et Biophysica Acta (BBA) - Biomembranes Volume: 1 Issue: 12 Pages: 183154-183154
  • DOI: 10.1016/j.bbamem.2019.183154
  • Peer Reviewed / Open Access
• [Journal Article] Structural Basis of H+-Dependent Conformational Change in a Bacterial MATE Transporter (2019)
  • Author(s): Kusakizako Tsukasa、Claxton Derek P.、Tanaka Yoshiki、Maturana Andrs D.、Kuroda Teruo、Ishitani Ryuichiro、Mchaourab Hassane S.、Nureki Osamu
  • Journal Title: Structure Volume: 27 Issue: 2 Pages: 293-301
  • DOI: 10.1016/j.str.2018.10.004
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Crystal structure of the Agrobacterium tumefaciens type VI effector-immunity complex. (2018)
  • Author(s): Fukuhara S, Nakane T, Yamashita K, Ishii R, Ishitani R, Nureki O
  • Journal Title: Acta Crystallogr. F Struct. Biol. Commun. Volume: 74 Issue: 3 Pages: 810-816
  • DOI: 10.1038/s41477-019-0367-2
  • Peer Reviewed / Open Access
• [Journal Article] Cap-specific terminal N6-methylation of RNA by an RNA polymerase II?associated methyltransferase (2018)
  • Author(s): Akichika Shinichiro、Hirano Seiichi、Shichino Yuichi、Suzuki Takeo、Nishimasu Hiroshi、Ishitani Ryuichiro、Sugita Ai、Hirose Yutaka、Iwasaki Shintaro、Nureki Osamu、Suzuki Tsutomu
  • Journal Title: Science Volume: 363 Issue: 363 Pages: 1-7
  • DOI: 10.1126/science.aav0080
  • Peer Reviewed / Open Access
• [Journal Article] Structural insights into cGAMP degradation by Ecto-nucleotide pyrophosphatase phosphodiesterase 1 (2018)
  • Author(s): Kato Kazuki、Nishimasu Hiroshi、Oikawa Daisuke、Hirano Seiichi、Hirano Hisato、Kasuya Go、Ishitani Ryuichiro、Tokunaga Fuminori、Nureki Osamu
  • Journal Title: Nature Communications Volume: 9 Issue: 1 Pages: 4424-4424
  • DOI: 10.1038/s41467-018-06922-7
  • Peer Reviewed / Open Access
• [Journal Article] Vibrational and Molecular Properties of Mg2+ Binding and Ion Selectivity in the Magnesium Channel MgtE (2018)
  • Author(s): Kimura Tetsunari、Lorenz-Fonfria Victor A.、Douki Shintaro、Motoki Hideyoshi、Ishitani Ryuichiro、Nureki Osamu、Higashi Masahiro、Furutani Yuji
  • Journal Title: The Journal of Physical Chemistry B Volume: 122 Issue: 42 Pages: 9681-9696
  • DOI: 10.1021/acs.jpcb.8b07967
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] An Atomistic Model of a Precursor State of Light-Induced Channel Opening of Channelrhodopsin (2018)
  • Author(s): Cheng Cheng, Motoshi Kamiya, Mizuki Takemoto, Ryuichiro Ishitani, Osamu Nureki, *Norio Yoshida, and *Shigehiko Hayashi
  • Journal Title: Biophysical Journal Volume: 115 Issue: 7 Pages: 1281-1291
  • DOI: 10.1016/j.bpj.2018.08.024
  • Peer Reviewed / Open Access
• [Journal Article] Free Energy Landscape for the Entire Transport Cycle of Triose-Phosphate/Phosphate Translocator (2018)
  • Author(s): Takemoto M, Lee Y, Ishitani R, Nureki O
  • Journal Title: Structure Volume: 26 Issue: 9 Pages: 1284-1296
  • DOI: 10.1016/j.str.2018.05.012
  • Peer Reviewed / Open Access
• [Journal Article] Engineered CRISPR-Cas9 nuclease with expanded targeting space (2018)
  • Author(s): Nishimasu Hiroshi、Shi Xi、Ishiguro Soh、Gao Linyi、Hirano Seiichi、Okazaki Sae、Noda Taichi、Abudayyeh Omar O.、Gootenberg Jonathan S.、Mori Hideto、Oura Seiya、Holmes Benjamin、Tanaka Mamoru、Seki Motoaki、Hirano Hisato、Aburatani Hiroyuki、Ishitani Ryuichiro、Ikawa Masahito、Yachie Nozomu、Zhang Feng、Nureki Osamu
  • Journal Title: Science Volume: 361 Issue: 6408 Pages: 1259-1262
  • DOI: 10.1126/science.aas9129
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Cryo-EM structures of the human volume-regulated anion channel LRRC8 (2018)
  • Author(s): Kasuya G, Nakane T, Yokoyama T, Jia Y, Inoue M, Watanabe K, Nakamura R, Nishizawa T, Kusakizako T, Tsutsumi A, Yanagisawa H, Dohmae N, Hattori M, Ichijo H, Yan Z, Kikkawa M, Shirouzu M, Ishitani R, Nureki O
  • Journal Title: Nat Struct Mol Biol. Volume: 25 Issue: 9 Pages: 797-804
  • DOI: 10.1038/s41594-018-0109-6
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Functional roles of Mg2+ binding sites in ion-dependent gating of a Mg2+ channel, MgtE, revealed by solution NMR (2018)
  • Author(s): Maruyama Tatsuro、Imai Shunsuke、Kusakizako Tsukasa、Hattori Motoyuki、Ishitani Ryuichiro、Nureki Osamu、Ito Koichi、Maturana Andr?s D、Shimada Ichio、Osawa Masanori
  • Journal Title: eLife Volume: 7
  • DOI: 10.7554/elife.31596
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural basis for xenobiotics extrusion by eukaryotic MATE transporter. (2017)
  • Author(s): Miyauchi H., Moriyama S., Kusakizako T., Kumazaki K., Ito K., Dohmae N., Nishizawa T., Miyaji T., Moriyama Y., Ishitani R., Nureki O.
  • Journal Title: Nature Commun. Volume: 8 Issue: 1 Pages: 1633-1633
  • DOI: 10.1038/s41467-017-01541-0
  • Peer Reviewed / Open Access
• [Journal Article] Structural insights into the competitive inhibition of the ATP-gated P2X receptor channel (2017)
  • Author(s): Kasuya G, Yamaura T, Ma XB, Nakamura R, Takemoto M, Nagumo H, Tanaka E, Dohmae N, Nakane T, Yu Y, Ishitani R, Matsuzaki O, Hattori M, Nureki O
  • Journal Title: Nat Commun. Volume: 8 Issue: 1 Pages: 876-876
  • DOI: 10.1038/s41467-017-00887-9
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structure of the triose-phosphate/phosphate translocator reveals the basis of substrate specificity. (2017)
  • Author(s): Lee Y, Nishizawa T, Takemoto M, Kumazaki K, Yamashita K, Hirata K, Minoda A, Nagatoishi S, Tsumoto K, Ishitani R and Nureki O.
  • Journal Title: Nature Plants. Volume: 3 Issue: 10 Pages: 825-832
  • DOI: 10.1038/s41477-017-0022-8
  • Peer Reviewed / Open Access
• [Journal Article] Structural insights into ligand recognition by the lysophosphatidic acid receptor LPA6 (2017)
  • Author(s): Taniguchi Reiya、Inoue Asuka、Sayama Misa、Uwamizu Akiharu、Yamashita Keitaro、Hirata Kunio、Yoshida Masahito、Tanaka Yoshiki、Kato Hideaki E.、Nakada-Nakura Yoshiko、Otani Yuko、Nishizawa Tomohiro、Doi Takayuki、Ohwada Tomohiko、Ishitani Ryuichiro、Aoki Junken、Nureki Osamu
  • Journal Title: Nature Volume: 548 Issue: 7667 Pages: 356-360
  • DOI: 10.1038/nature23448
  • Peer Reviewed / Open Access
• [Journal Article] Structural Basis for the Canonical and Non-canonical PAM Recognition by CRISPR-Cpf1 (2017)
  • Author(s): Yamano Takashi、Zetsche Bernd、Ishitani Ryuichiro、Zhang Feng、Nishimasu Hiroshi、Nureki Osamu
  • Journal Title: Molecular Cell Volume: 67 Issue: 4 Pages: 633-645
  • DOI: 10.1016/j.molcel.2017.06.035
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] ATP-dependent modulation of MgtE in Mg2+ homeostasis (2017)
  • Author(s): Tomita Atsuhiro、Zhang Mingfeng、Jin Fei、Zhuang Wenhui、Takeda Hironori、Maruyama Tatsuro、Osawa Masanori、Hashimoto Ken-ichi、Kawasaki Hisashi、Ito Koichi、Dohmae Naoshi、Ishitani Ryuichiro、Shimada Ichio、Yan Zhiqiang、Hattori Motoyuki、Nureki Osamu
  • Journal Title: Nature Communications Volume: 8 Issue: 1 Pages: 148-148
  • DOI: 10.1038/s41467-017-00082-w
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Structural Basis for the Altered PAM Recognition by Engineered CRISPR-Cpf1 (2017)
  • Author(s): Nishimasu Hiroshi、Yamano Takashi、Gao Linyi、Zhang Feng、Ishitani Ryuichiro、Nureki Osamu
  • Journal Title: Molecular Cell Volume: 67 Issue: 1 Pages: 139-147
  • DOI: 10.1016/j.molcel.2017.04.019
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Cyclic GMP-AMP as an Endogenous Second Messenger in Innate Immune Signaling by Cytosolic DNA (2017)
  • Author(s): Kato K, Omura H, Ishitani R, Nureki O
  • Journal Title: Annu Rev Biochem Volume: 86 Issue: 1 Pages: 541-566
  • DOI: 10.1146/annurev-biochem-061516-044813
  • Peer Reviewed / Open Access
• [Journal Article] Structural insights into the nucleotide base specificity of P2X receptors. (2017)
  • Author(s): Kasuya G, Fujiwara Y, Tsukamoto H, Morinaga S, Ryu S, Touhara K, Ishitani R, Furutani Y, Hattori M, Nureki O.
  • Journal Title: Sci Rep. Volume: 7 Issue: 1 Pages: 45208-45208
  • DOI: 10.1038/srep45208
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Crystal Structure of the Minimal Cas9 from Campylobacter jejuni Reveals the Molecular Diversity in the CRISPR-Cas9 Systems (2016)
  • Author(s): Yamada M, Watanabe Y, Gootenberg JS, Hirano H, Ran FA, Nakane T, Ishitani R, Zhang F, Nishimasu H, Nureki O
  • Journal Title: Mol Cell Volume: 65 Issue: 6 Pages: 1109-1121
  • DOI: 10.1016/j.molcel.2017.02.007
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant