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Elucidation of effective consecutive chemical reaction in cell with formation of supramolecular enzyme complex

Research Project

Project/Area Number 17H03092
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Bio-related chemistry
Research InstitutionInstitute of Physical and Chemical Research

Principal Investigator

Tosha Takehiko  国立研究開発法人理化学研究所, 放射光科学研究センター, 専任研究員 (00548993)

Co-Investigator(Kenkyū-buntansha) 新井 博之  東京大学, 大学院農学生命科学研究科(農学部), 准教授 (70291052)
Project Period (FY) 2017-04-01 – 2021-03-31
Project Status Completed (Fiscal Year 2020)
Budget Amount *help
¥18,330,000 (Direct Cost: ¥14,100,000、Indirect Cost: ¥4,230,000)
Fiscal Year 2020: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
Fiscal Year 2019: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
Fiscal Year 2018: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
Fiscal Year 2017: ¥10,920,000 (Direct Cost: ¥8,400,000、Indirect Cost: ¥2,520,000)
Keywords蛋白質複合体 / 一酸化窒素 / 脱窒 / 金属酵素 / 反応機構 / 時間分解計測 / 超分子複合体 / ヘム / 相互作用 / 酵素反応 / 電子顕微鏡 / CryoEM / タンパク質複合体 / 金属タンパク質
Outline of Final Research Achievements

The molecular mechanism in bacterial denitrification, a kind of anaerobic respiration, consisting of four consecutive chemical reactions was investigated to elucidate how biological system effectively conducts sequential reaction in cells. We found that, to minimize the cytotoxicity of nitric oxide (NO) which is an intermediate product in denitrification, NO-producing enzyme, nitrite reductase (NiR) and NO-decomposing enzyme, NO reductase (NOR), forms a complex, and NOR can decompose NO immediately after its production from nitrite by NiR. In addition, the reaction mechanism of the rapid NO decomposition by NOR was examined. Time-resolved visible absorption spectroscopic analysis using photosensitive caged NO as a reaction trigger showed that two NO molecules bind to the active site of NOR with step-wise manner for NO decomposition.

Academic Significance and Societal Importance of the Research Achievements

細胞内は、生体分子でみたされたクラウディングという状態にあり、生体内での反応は、多数の夾雑物が存在する条件下で行われる。脱窒のように連続した化学反応が分子夾雑下で効率よく行われるためには、それぞれの反応を触媒する酵素タンパク質が連携して強調的に機能する仕組みが必要となる。このような問いに対する一つの回答として、本研究から、脱窒に関わるタンパク質が相互作用することで、中間生成物を拡散させずに効率よく逐次反応を行っていることが示唆された。一連の生理反応に関わるタンパク質が相互作用して機能するという概念は、脱窒以外の反応系にも適用できる考え方であり、一般性のある現象であることが推察される。

Report

(5 results)
  • 2020 Annual Research Report   Final Research Report ( PDF )
  • 2019 Annual Research Report
  • 2018 Annual Research Report
  • 2017 Annual Research Report
  • Research Products

    (50 results)

All 2021 2020 2019 2018 2017 Other

All Int'l Joint Research (5 results) Journal Article (18 results) (of which Int'l Joint Research: 6 results,  Peer Reviewed: 18 results,  Open Access: 12 results) Presentation (23 results) (of which Int'l Joint Research: 8 results,  Invited: 13 results) Book (1 results) Remarks (3 results)

  • [Int'l Joint Research] University of Essex/University of Liverpool/Diamond Light Source(英国)

    • Related Report
      2020 Annual Research Report
  • [Int'l Joint Research] University of Liverpool/University of Leeds(英国)

    • Related Report
      2019 Annual Research Report
  • [Int'l Joint Research] リバプール大学(英国)

    • Related Report
      2018 Annual Research Report
  • [Int'l Joint Research] リバプール大学(英国)

    • Related Report
      2017 Annual Research Report
  • [Int'l Joint Research] ストックホルム大学(スウェーデン)

    • Related Report
      2017 Annual Research Report
  • [Journal Article] An unprecedented insight into the catalytic mechanism of copper nitrite reductase from atomic-resolution and damage-free structures2021

    • Author(s)
      Rose Samuel L.、Antonyuk Svetlana V.、Sasaki Daisuke、Yamashita Keitaro、Hirata Kunio、Ueno Go、Ago Hideo、Eady Robert R.、Tosha Takehiko、Yamamoto Masaki、Hasnain S. Samar
    • Journal Title

      Science Advances

      Volume: 7 Issue: 1

    • DOI

      10.1126/sciadv.abd8523

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Impact of membrane protein-lipid interactions on formation of bilayer lipid membranes on SAM-modified gold electrode2021

    • Author(s)
      Kato Masaru、Masuda Yuya、Yoshida Narumi、Tosha Takehiko、Shiro Yoshitsugu、Yagi Ichizo
    • Journal Title

      Electrochimica Acta

      Volume: 373 Pages: 137888-137888

    • DOI

      10.1016/j.electacta.2021.137888

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] NO Dynamics in Microbial Denitrification System2021

    • Author(s)
      Tosha Takehiko、Yamagiwa Raika、Sawai Hitomi、Shiro Yoshitsugu
    • Journal Title

      Chemistry Letters

      Volume: 50 Issue: 2 Pages: 280-288

    • DOI

      10.1246/cl.200629

    • NAID

      130007986360

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Timing of NO Binding and Protonation in the Catalytic Reaction of Bacterial Nitric Oxide Reductase as Established by Time-Resolved Spectroscopy.2020

    • Author(s)
      Hanae Takeda, Tatsunari Kimura, Takashi Nomura, Masaki Horitani, Azusa Yokota, Akiko Matsubayashi, Shoko Ishii, Yoshitsugu Shiro, Minoru Kubo and Takehiko Tosha
    • Journal Title

      Bulletin of the Chemical Society of Japan

      Volume: - Issue: 7 Pages: 825-833

    • DOI

      10.1246/bcsj.20200038

    • NAID

      130007879554

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Serial Femtosecond Zero Dose Crystallography Captures a Water‐Free Distal Heme Site in a Dye‐Decolorising Peroxidase to Reveal a Catalytic Role for an Arginine in Fe IV =O Formation2020

    • Author(s)
      Lucic Marina、Svistunenko Dimitri A.、Wilson Michael T.、Chaplin Amanda K.、Davy Bradley、Ebrahim Ali、Axford Danny、Tosha Takehiko、Sugimoto Hiroshi、Owada Shigeki、Dworkowski Florian S. N.、Tews Ivo、Owen Robin L.、Hough Michael A.、Worrall Jonathan A. R.
    • Journal Title

      Angewandte Chemie International Edition

      Volume: 59 Issue: 48 Pages: 21656-21662

    • DOI

      10.1002/anie.202008622

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Time-resolved studies of metalloproteins using X-ray free electron laser radiation at SACLA2020

    • Author(s)
      Suga Michihiro、Shimada Atsuhiro、Akita Fusamichi、Shen Jian-Ren、Tosha Takehiko、Sugimoto Hiroshi
    • Journal Title

      Biochimica et Biophysica Acta (BBA) - General Subjects

      Volume: 1864 Issue: 2 Pages: 129466-129466

    • DOI

      10.1016/j.bbagen.2019.129466

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Hijacking the Heme Acquisition System of Pseudomonas aeruginosa for the Delivery of Phthalocyanine as an Antimicrobial2019

    • Author(s)
      Shisaka Yuma、Iwai Yusuke、Yamada Shiho、Uehara Hiromu、Tosha Takehiko、Sugimoto Hiroshi、Shiro Yoshitsugu、Stanfield Joshua K.、Ogawa Kazuya、Watanabe Yoshihito、Shoji Osami
    • Journal Title

      ACS Chemical Biology

      Volume: 14 Issue: 7 Pages: 1637-1642

    • DOI

      10.1021/acschembio.9b00373

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Dimeric structures of quinol-dependent nitric oxide reductases (qNORs) revealed by cryo-electron microscopy2019

    • Author(s)
      Gopalasingam Chai C.、Johnson Rachel M.、Chiduza George N.、Tosha Takehiko、Yamamoto Masaki、Shiro Yoshitsugu、Antonyuk Svetlana V.、Muench Stephen P.、Hasnain S. Samar
    • Journal Title

      Science Advances

      Volume: 5 Issue: 8

    • DOI

      10.1126/sciadv.aax1803

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Visualization of Enzymatic Reactions Using SACLA2019

    • Author(s)
      TOSHA Takehiko、KUBO Minoru
    • Journal Title

      Seibutsu Butsuri

      Volume: 59 Issue: 4 Pages: 205-207

    • DOI

      10.2142/biophys.59.205

    • NAID

      130007683542

    • ISSN
      0582-4052, 1347-4219
    • Related Report
      2019 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Structural basis for promotion of duodenal iron absorption by enteric ferric reductase with ascorbate.2018

    • Author(s)
      Ganasen M, Togashi H, Takeda H, Asakura H, Tosha T, Yamashita K, Hirata K, Nariai Y, Urano T, Yuan X, Hamza I, Mauk AG, Shiro Y, Sugimoto H, Sawai H.
    • Journal Title

      Communications Biology

      Volume: 1 Issue: 1 Pages: 120-120

    • DOI

      10.1038/s42003-018-0121-8

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Identification of a novel zinc-binding protein, C1orf123, as an interactor with a heavy metal-associated domain2018

    • Author(s)
      Furukawa Yoshiaki、Lim Carolyn、Tosha Takehiko、Yoshida Koki、Hagai Tomoaki、Akiyama Shuji、Watanabe Shoji、Nakagome Kenta、Shiro Yoshitsugu
    • Journal Title

      PLOS ONE

      Volume: 13 Issue: 9 Pages: e0204355-e0204355

    • DOI

      10.1371/journal.pone.0204355

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Surface-Enhanced Infrared Absorption Spectroscopy of Bacterial Nitric Oxide Reductase under Electrochemical Control Using Vibrational Probe of Carbon Monoxide2018

    • Author(s)
      M. Kato, S. Nakagawa, T. Tosha, Y. Shiro, Y. Masuda, K. Nakata, I. Yagi
    • Journal Title

      J. Phys. Chem. Lett.

      Volume: 9 Issue: 17 Pages: 5196-5200

    • DOI

      10.1021/acs.jpclett.8b02581

    • NAID

      120006887743

    • Related Report
      2018 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Pseudomonas aeruginosa overexpression system of nitric oxide reductase for in vivo and in vitro mutational analyses2018

    • Author(s)
      Yamagiwa Raika、Kurahashi Takuya、Takeda Mariko、Adachi Mayuho、Nakamura Hiro、Arai Hiroyuki、Shiro Yoshitsugu、Sawai Hitomi、Tosha Takehiko
    • Journal Title

      Biochimica et Biophysica Acta (BBA) - Bioenergetics

      Volume: 1859 Issue: 5 Pages: 333-341

    • DOI

      10.1016/j.bbabio.2018.02.009

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Characterization of the Quinol-dependent Nitric Oxide Reductase from the Pathogen Neisseria meningitidis, an Electrogenic Enzyme2018

    • Author(s)
      N. Gonska, D. R. Young, R. Yuki, T. Okamoto, T. Hisano, S. V. Antonyuk, S. S. Hasnain, K. Muramoto, Y. Shiro, T. Tosha, P. Adelroth
    • Journal Title

      Sci. Rep.

      Volume: 8: 3637 Issue: 1 Pages: 3637-3637

    • DOI

      10.1038/s41598-018-21804-0

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] An unprecedented dioxygen species revealed by serial femtosecond rotation crystallography in copper nitrite reductase2018

    • Author(s)
      Halsted Thomas P.、Yamashita Keitaro、Hirata Kunio、Ago Hideo、Ueno Go、Tosha Takehiko、Eady Robert R.、Antonyuk Svetlana V.、Yamamoto Masaki、Hasnain S. Samar
    • Journal Title

      IUCrJ

      Volume: 5 Issue: 1 Pages: 22-31

    • DOI

      10.1107/s2052252517016128

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Capturing an initial intermediate during the P450nor enzymatic reaction using time-resolved XFEL crystallography and caged-substrate2017

    • Author(s)
      Tosha Takehiko、Kubo Minoru, et. al
    • Journal Title

      Nature Communications

      Volume: 8 Issue: 1 Pages: 1-9

    • DOI

      10.1038/s41467-017-01702-1

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Structural basis for binding and transfer of heme in bacterial heme-acquisition systems2017

    • Author(s)
      Naoe Youichi、Nakamura Nozomi、Rahman Md. Mahfuzur、Tosha Takehiko、Nagatoishi Satoru、Tsumoto Kouhei、Shiro Yoshitsugu、Sugimoto Hiroshi
    • Journal Title

      Proteins: Structure, Function, and Bioinformatics

      Volume: 85 Issue: 12 Pages: 2217-2230

    • DOI

      10.1002/prot.25386

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Dynamics of Nitric Oxide Controlled by Protein Complex in Bacterial System2017

    • Author(s)
      Erina Terasaka, Kenta Yamada, Po-Hung Wang, Kanta Hosokawa, Raika Yamagiwa, Kimi Matsumoto, Shoko Ishii, Takaharu Mori, Kiyoshi Yagi, Hitomi Sawai, Hiroyuki Arai, Hiroshi Sugimoto, Yuji Sugita, Yoshitsugu Shiro and Takehiko Tosha
    • Journal Title

      PNAS

      Volume: 114 Issue: 37 Pages: 9888-9893

    • DOI

      10.1073/pnas.1621301114

    • Related Report
      2017 Annual Research Report
    • Peer Reviewed
  • [Presentation] 光解離性ケージド基質を利用した時間分解構造解析による酵素反応の可視化2021

    • Author(s)
      當舎武彦
    • Organizer
      日本薬学会第141回年会
    • Related Report
      2020 Annual Research Report
    • Invited
  • [Presentation] 構造生物学におけるXFELと放射光の相補的利用~金属酵素の反応機構解明を目指して~2021

    • Author(s)
      當舎武彦
    • Organizer
      放射光ユーザーのためのSACLAの利活用に関するワークショップ
    • Related Report
      2020 Annual Research Report
    • Invited
  • [Presentation] マイクロ秒時間領域で形成される一酸化窒素還元酵素反応中間体の分光解析2020

    • Author(s)
      當舎武彦
    • Organizer
      第58回日本生物物理学会年会
    • Related Report
      2020 Annual Research Report
  • [Presentation] 時間分解結晶構造解析および分光解析が明らかにする金属酵素による一酸化窒素分解機構2020

    • Author(s)
      當舎武彦
    • Organizer
      第2回量子線科学セミナー
    • Related Report
      2020 Annual Research Report
    • Invited
  • [Presentation] Elucidation of NO reduction mechanism in soluble NO reductase by time-resolved crystallography with photosensitive caged compound2019

    • Author(s)
      Takehiko Tosha
    • Organizer
      32nd European Crystallographic Meeting (ECM32)
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] Mechanism of P450nor-Catalyzed NO Reduction Proved by Time-Resolved Spectroscopic and Crystallographic Analyses2019

    • Author(s)
      T. Tosha, T. Nomura, H. Takeda, T. Kimura, H. Sugimoto, M. Kubo and Y. Shiro
    • Organizer
      19th International Conference on Biological Inorganic Chemistry
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Functional roles of conserved residues near the active site of nitric oxide reductase based on the structural analysis2019

    • Author(s)
      當舎武彦、山際来佳、倉橋拓也、新井博之、杉本宏、城宜嗣
    • Organizer
      第57回日本生物物理学会年会
    • Related Report
      2019 Annual Research Report
  • [Presentation] NO-binding and protonation process in the catalytic reaction of the bacterial Nitric oxide reductase as established by time-resolved spectroscopy2019

    • Author(s)
      武田英恵、木村哲就、野村高志、當舎武彦、城宜嗣、久保稔
    • Organizer
      第57回日本生物物理学会年会
    • Related Report
      2019 Annual Research Report
  • [Presentation] NO-binding and Protonation Process in the Catalytic Reaction of Heme/non-heme Iron Nitric Oxide Reductase Proved by Time-Resolved Spectroscopic System2019

    • Author(s)
      H. Takeda, T. Kimura, T. Nomura, T. Tosha, Y. Shiro, M. Kubo
    • Organizer
      15th International Symposium on Applied Bioinorganic Chemistry(ISABIC)
    • Related Report
      2019 Annual Research Report
    • Int'l Joint Research
  • [Presentation] NITRIC OXIDE DYNAMICS CONTROLLED BY FORMATION OF PROTEIN COMPLEX IN DENTRIFICATION2018

    • Author(s)
      Takehiko Tosha, Erina Terasaka, Kenta Yamada, Po-Hung Wang, Hiroyuki Arai, Hiroshi Sugimoto, Yuji Sugita, Yoshistugu Shiro
    • Organizer
      9th Asian Biological Inorganic Chemistry Conference
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] Regulation of nitric oxide dynamics in microbial denitrification2018

    • Author(s)
      Takehiko Tosha
    • Organizer
      The International Symposium on Bioinorganic Chemistry 2018
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] Elucidation of Mechanism of Biological Nitric Oxide Reduction2018

    • Author(s)
      Takehiko Tosha, Hanae Takeda, Takashi Nomura, Tetsunari Kimura, Minoru Kubo, Yoshitsugu Shiro
    • Organizer
      43rd International Conference on Coordination Chemistry
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Mechanism of Biological Nitric Oxide Reduction Proved by Time-resolved Spectroscopic Analyses2018

    • Author(s)
      Tosha Tosha, Tetsunari Kimura, Hanae Takeda, Shoko Ishii, Takashi Nomura, Minoru Kubo, Yoshitsugu Shiro
    • Organizer
      International Conference on Porphyrins and Phthalocyanines
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] 亜硝酸還元酵素と一酸化窒素還元酵素の複合体形成による迅速なNO分解機構2018

    • Author(s)
      當舎武彦
    • Organizer
      平成30年度 内外環境応答・代謝酵素研究会
    • Related Report
      2018 Annual Research Report
  • [Presentation] 脱窒にみられる金属タンパク質複合体による効率的な連続化学反応2018

    • Author(s)
      當舎武彦
    • Organizer
      第56回日本生物物理学会年会
    • Related Report
      2018 Annual Research Report
    • Invited
  • [Presentation] Mechanism of Biological Nitric Oxide Reduction2018

    • Author(s)
      當舎武彦
    • Organizer
      平成30年度化学系学協会東北大会
    • Related Report
      2018 Annual Research Report
    • Invited
  • [Presentation] 光解離性基質を利用した時間分解X線結晶構造解析による酵素反応の追跡2018

    • Author(s)
      當舎武彦
    • Organizer
      第18回日本蛋白質科学会年会
    • Related Report
      2018 Annual Research Report
    • Invited
  • [Presentation] Time-resolved XFEL crystallography and in crystallo spectroscopy for probing reaction dynamics of respiratory metalloenzymes2018

    • Author(s)
      當舎武彦
    • Organizer
      5th BioXFEL International Conference
    • Related Report
      2017 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] X線自由電子レーザーを用いた時間分解結晶構造解析による酵素反応の追跡2018

    • Author(s)
      當舎武彦
    • Organizer
      日本化学会第98春季年会
    • Related Report
      2017 Annual Research Report
    • Invited
  • [Presentation] 酵素反応機構解明のためのX線自由電子レーザーを利用した時間分解X線結晶構造解析2017

    • Author(s)
      當舎武彦、西田拓真、野村高志、佐伯直哉、杉本宏、山下恵太郎、平田邦生、吾郷日出夫、山本雅貴、中根崇智、菅原道泰、南後恵理子、岩田想、城宜嗣、久保稔
    • Organizer
      第50回酸化反応討論会
    • Related Report
      2017 Annual Research Report
  • [Presentation] X線自由電子レーザーを用いた時間分解結晶構造解析:酵素反応への応用2017

    • Author(s)
      當舎武彦
    • Organizer
      第55回日本生物物理学会年会
    • Related Report
      2017 Annual Research Report
    • Invited
  • [Presentation] Time-resolved X-ray crystallography at SACLA: Application to enzymatic reaction2017

    • Author(s)
      當舎武彦、西田拓真、野村高志、佐伯直哉、杉本宏、山下恵太郎、平田邦生、吾郷日出夫、山本雅貴、中根崇智、菅原道泰、南後恵理子、岩田想、城宜嗣、久保稔
    • Organizer
      錯体化学会第67回討論会
    • Related Report
      2017 Annual Research Report
  • [Presentation] X線自由電子レーザーを利用した時間分解X線結晶構造解析による酵素反応の観測2017

    • Author(s)
      當舎武彦、西田拓真、野村高志、佐伯直哉、杉本宏、山下恵太郎、平田邦生、吾郷日出夫、山本雅貴、中根崇智、菅原道泰、南後恵理子、岩田想、城宜嗣、久保稔
    • Organizer
      第44回生体分子科学討論会
    • Related Report
      2017 Annual Research Report
  • [Book] Dioxygen-dependent heme enzymes2019

    • Author(s)
      Masao Ikeda-Saito, Emma Raven
    • Total Pages
      376
    • Publisher
      Royal Society of Chemistry
    • Related Report
      2018 Annual Research Report
  • [Remarks] SACLAで酵素反応の可視化を実現

    • URL

      http://www.riken.jp/pr/press/2017/20171117_2/

    • Related Report
      2017 Annual Research Report
  • [Remarks] 合成酵素と分解酵素の協演

    • URL

      http://www.riken.jp/pr/press/2017/20170829_1/

    • Related Report
      2017 Annual Research Report
  • [Remarks] 病原菌における抗菌ガス分解酵素の新機能

    • URL

      http://www.riken.jp/pr/press/2018/20180227_1/

    • Related Report
      2017 Annual Research Report

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Published: 2017-04-28   Modified: 2022-01-27  

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