Performance information

• A multipoint guidance mechanism for β-barrel folding on the SAM complex　　　　　　　　　　　　　　　
  Hironori Takeda; Jon V. Busto; Caroline Lindau; Akihisa Tsutsumi; Kentaro Tomii; Kenichiro Imai; Yu Yamamori; Takatsugu Hirokawa; Chie Motono; Iniyan Ganesan; Lena-Sophie Wenz; Thomas Becker; Masahide Kikkawa; Nikolaus Pfanner; Nils Wiedemann; Toshiya Endo
  Nature Structural and Molecular Biology, Volume：30, Number：2, First page：176, Last page：187, Jan. 2023
  Springer Science and Business Media LLC, Scientific journal
  DOI：https://doi.org/10.1038/s41594-022-00897-2
  DOI ID：10.1038/s41594-022-00897-2, ISSN：1545-9993, eISSN：1545-9985, Web of Science ID：WOS:000909509300001
• The structure of MgtE in the absence of magnesium provides new insights into channel gating
  Fei Jin; Minxuan Sun; Takashi Fujii; Yurika Yamada; Jin Wang; Andrés D. Maturana; Miki Wada; Shichen Su; Jinbiao Ma; Hironori Takeda; Tsukasa Kusakizako; Atsuhiro Tomita; Yoshiko Nakada-Nakura; Kehong Liu; Tomoko Uemura; Yayoi Nomura; Norimichi Nomura; Koichi Ito; Osamu Nureki; Keiichi Namba; So Iwata; Ye Yu; Motoyuki Hattori
  PLOS Biology, Volume：19, Number：4, First page：e3001231, Last page：e3001231, Apr. 2021
  MgtE is a Mg²⁺ channel conserved in organisms ranging from prokaryotes to eukaryotes, including humans, and plays an important role in Mg²⁺ homeostasis. The previously determined MgtE structures in the Mg²⁺-bound, closed-state, and structure-based functional analyses of MgtE revealed that the binding of Mg²⁺ ions to the MgtE cytoplasmic domain induces channel inactivation to maintain Mg²⁺ homeostasis. There are no structures of the transmembrane (TM) domain for MgtE in Mg²⁺-free conditions, and the pore-opening mechanism has thus remained unclear.
  
  Here, we determined the cryo-electron microscopy (cryo-EM) structure of the MgtE-Fab complex in the absence of Mg²⁺ ions. The Mg²⁺-free MgtE TM domain structure and its comparison with the Mg²⁺-bound, closed-state structure, together with functional analyses, showed the Mg²⁺-dependent pore opening of MgtE on the cytoplasmic side and revealed the kink motions of the TM2 and TM5 helices at the glycine residues, which are important for channel activity. Overall, our work provides structure-based mechanistic insights into the channel gating of MgtE.
  Public Library of Science (PLoS), Scientific journal
  DOI：https://doi.org/10.1371/journal.pbio.3001231
  DOI ID：10.1371/journal.pbio.3001231, eISSN：1545-7885
• Mitochondrial sorting and assembly machinery operates by β-barrel switching.　　　　　　　　　　　　　　　
  Hironori Takeda; Akihisa Tsutsumi; Tomohiro Nishizawa; Caroline Lindau; Jon V Busto; Lena-Sophie Wenz; Lars Ellenrieder; Kenichiro Imai; Sebastian P Straub; Waltraut Mossmann; Jian Qiu; Yu Yamamori; Kentaro Tomii; Junko Suzuki; Takeshi Murata; Satoshi Ogasawara; Osamu Nureki; Thomas Becker; Nikolaus Pfanner; Nils Wiedemann; Masahide Kikkawa; Toshiya Endo
  Nature, Volume：590, Number：7844, First page：163, Last page：169, Feb. 2021, ［International magazine］
  The mitochondrial outer membrane contains so-called β-barrel proteins, which allow communication between the cytosol and the mitochondrial interior1-3. Insertion of β-barrel proteins into the outer membrane is mediated by the multisubunit mitochondrial sorting and assembly machinery (SAM, also known as TOB)4-6. Here we use cryo-electron microscopy to determine the structures of two different forms of the yeast SAM complex at a resolution of 2.8-3.2 Å. The dimeric complex contains two copies of the β-barrel channel protein Sam50-Sam50a and Sam50b-with partially open lateral gates. The peripheral membrane proteins Sam35 and Sam37 cap the Sam50 channels from the cytosolic side, and are crucial for the structural and functional integrity of the dimeric complex. In the second complex, Sam50b is replaced by the β-barrel protein Mdm10. In cooperation with Sam50a, Sam37 recruits and traps Mdm10 by penetrating the interior of its laterally closed β-barrel from the cytosolic side. The substrate-loaded SAM complex contains one each of Sam50, Sam35 and Sam37, but neither Mdm10 nor a second Sam50, suggesting that Mdm10 and Sam50b function as placeholders for a β-barrel substrate released from Sam50a. Our proposed mechanism for dynamic switching of β-barrel subunits and substrate explains how entire precursor proteins can fold in association with the mitochondrial machinery for β-barrel assembly.
  English, Scientific journal
  DOI：https://doi.org/10.1038/s41586-020-03113-7
  DOI ID：10.1038/s41586-020-03113-7, PubMed ID：33408415
• Membrane Protein Insertion Mechanism by Mitochondrial Sorting and Assembly Machinery Complex
  Hironori TAKEDA; Toshiya ENDO
  Seibutsu Butsuri, Volume：61, Number：6, First page：392, Last page：394, 2021, [Reviewed], [Invited]
  Biophysical Society of Japan, Japanese, Scientific journal
  DOI：https://doi.org/10.2142/biophys.61.392
  DOI ID：10.2142/biophys.61.392, ISSN：0582-4052, eISSN：1347-4219
• ATP-dependent modulation of MgtE in Mg2+ homeostasis
  Atsuhiro Tomita; Mingfeng Zhang; Fei Jin; Wenhui Zhuang; Hironori Takeda; Tatsuro Maruyama; Masanori Osawa; Ken-ichi Hashimoto; Hisashi Kawasaki; Koichi Ito; Naoshi Dohmae; Ryuichiro Ishitani; Ichio Shimada; Zhiqiang Yan; Motoyuki Hattori; Osamu Nureki
  Nature Communications, Volume：8, Number：1, Jul. 2017
  Abstract
  
  Magnesium is an essential ion for numerous physiological processes. MgtE is a Mg²⁺ selective channel involved in the maintenance of intracellular Mg²⁺ homeostasis, whose gating is regulated by intracellular Mg²⁺ levels. Here, we report that ATP binds to MgtE, regulating its Mg²⁺-dependent gating. Crystal structures of MgtE–ATP complex show that ATP binds to the intracellular CBS domain of MgtE. Functional studies support that ATP binding to MgtE enhances the intracellular domain affinity for Mg²⁺ within physiological concentrations of this divalent cation, enabling MgtE to function as an in vivo Mg²⁺ sensor. ATP dissociation from MgtE upregulates Mg²⁺ influx at both high and low intracellular Mg²⁺ concentrations. Using site-directed mutagenesis and structure based-electrophysiological and biochemical analyses, we identify key residues and main structural changes involved in the process. This work provides the molecular basis of ATP-dependent modulation of MgtE in Mg²⁺ homeostasis.
  Springer Science and Business Media LLC, Scientific journal
  DOI：https://doi.org/10.1038/s41467-017-00082-w
  DOI ID：10.1038/s41467-017-00082-w, eISSN：2041-1723
• Structural basis for dynamic mechanism of nitrate/nitrite antiport by NarK.　　　　　　　　　　　　　　　
  Masahiro Fukuda; Hironori Takeda; Hideaki E Kato; Shintaro Doki; Koichi Ito; Andrés D Maturana; Ryuichiro Ishitani; Osamu Nureki
  Nature communications, Volume：6, First page：7097, Last page：7097, May 2015, ［International magazine］
  NarK belongs to the nitrate/nitrite porter (NNP) family in the major facilitator superfamily (MFS) and plays a central role in nitrate uptake across the membrane in diverse organisms, including archaea, bacteria, fungi and plants. Although previous studies provided insight into the overall structure and the substrate recognition of NarK, its molecular mechanism, including the driving force for nitrate transport, remained elusive. Here we demonstrate that NarK is a nitrate/nitrite antiporter, using an in vitro reconstituted system. Furthermore, we present the high-resolution crystal structures of NarK from Escherichia coli in the nitrate-bound occluded, nitrate-bound inward-open and apo inward-open states. The integrated structural, functional and computational analyses reveal the nitrate/nitrite antiport mechanism of NarK, in which substrate recognition is coupled to the transport cycle by the concomitant movement of the transmembrane helices and the key tyrosine and arginine residues in the substrate-binding site.
  English, Scientific journal
  DOI：https://doi.org/10.1038/ncomms8097
  DOI ID：10.1038/ncomms8097, PubMed ID：25959928, PubMed Central ID：PMC4432589
• Structural basis for ion selectivity revealed by high-resolution crystal structure of Mg2+ channel MgtE.　　　　　　　　　　　　　　　
  Hironori Takeda; Motoyuki Hattori; Tomohiro Nishizawa; Keitaro Yamashita; Syed T A Shah; Martin Caffrey; Andrés D Maturana; Ryuichiro Ishitani; Osamu Nureki
  Nature communications, Volume：5, First page：5374, Last page：5374, Nov. 2014, ［International magazine］
  Magnesium is the most abundant divalent cation in living cells and is crucial to several biological processes. MgtE is a Mg(2+) channel distributed in all domains of life that contributes to the maintenance of cellular Mg(2+) homeostasis. Here we report the high-resolution crystal structures of the transmembrane domain of MgtE, bound to Mg(2+), Mn(2+) and Ca(2+). The high-resolution Mg(2+)-bound crystal structure clearly visualized the hydrated Mg(2+) ion within its selectivity filter. Based on those structures and biochemical analyses, we propose a cation selectivity mechanism for MgtE in which the geometry of the hydration shell of the fully hydrated Mg(2+) ion is recognized by the side-chain carboxylate groups in the selectivity filter. This is in contrast to the K(+)-selective filter of KcsA, which recognizes a dehydrated K(+) ion. Our results further revealed a cation-binding site on the periplasmic side, which regulate channel opening and prevents conduction of near-cognate cations.
  English, Scientific journal
  DOI：https://doi.org/10.1038/ncomms6374
  DOI ID：10.1038/ncomms6374, PubMed ID：25367295, PubMed Central ID：PMC4241985

• Structural and functional study for mitohocndrial large channel formation.　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Early-Career Scientists, Apr. 2022 - Mar. 2025
  Nara Institute of Science and Technology
  Grant amount(Total)：4550000, Direct funding：3500000, Indirect funding：1050000
  Grant number：22K15078
• Optimization of multiple method baced on neutron reflectometry for analysis of biogenesis of Gram-negative bacterial membrane.　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Fund for the Promotion of Joint International Research (Fostering Joint International Research (B)), Fund for the Promotion of Joint International Research (Fostering Joint International Research (B)), Oct. 2021 - Mar. 2024
  University of Miyazaki
  Grant amount(Total)：18980000, Direct funding：14600000, Indirect funding：4380000
  Grant number：21KK0126
• Structural study for mitochondrial translocator SAM complex.　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Early-Career Scientists, Apr. 2018 - Mar. 2022
  Takeda Hironori
  Grant amount(Total)：4290000, Direct funding：3300000, Indirect funding：990000
  β-barrel membrane proteins localised on the mitochondrial outer membrane is essential for transport of small molecules and mitochondrial functions . The SAM complex embeds these β-barrel membrane proteins into the membrane. We determined the cryo-EM structures of the SAM complex and the SAM-Mdm10 supercomplex, and elucidated the substrate exchange mechanism by the SAM complex. This study was published in Nature. Moreover, we have successfully determined the cryo-EM structure of the SAM complex in the process of embedding the substrate protein into the membrane and also clarified a part of the molecular mechanism.
  Grant number：18K14640
• ミトコンドリアトランスロケーターＳＡＭ複合体の構造・機能研究　　　　　　　　　　　　　　　
  Apr. 2018 - Mar. 2021
  Grant amount(Total)：4030000, Direct funding：3100000, Indirect funding：930000
  Grant number：18J00358
• X線結晶構造解析を基盤としたマグネシウムイオン輸送体MgtEの開構造の解明　　　　　　　　　　　　　　　
  2013 - 2014
  Grant amount(Total)：1800000, Direct funding：1800000
  Grant number：13J07168