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Integrated structural analysis of clock protein KaiC in solution with small-angle scattering and analytical ultracentrifugation

Research Project

Project/Area Number 19K16088
Research Category

Grant-in-Aid for Early-Career Scientists

Allocation TypeMulti-year Fund
Review Section Basic Section 43040:Biophysics-related
Research InstitutionKyoto University

Principal Investigator

Morishima Ken  京都大学, 複合原子力科学研究所, 助教 (40812087)

Project Period (FY) 2019-04-01 – 2021-03-31
Project Status Completed (Fiscal Year 2020)
Budget Amount *help
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2020: ¥780,000 (Direct Cost: ¥600,000、Indirect Cost: ¥180,000)
Fiscal Year 2019: ¥3,380,000 (Direct Cost: ¥2,600,000、Indirect Cost: ¥780,000)
Keywords超遠心分析 (AUC) / X線小角散乱 (SAXS) / 時計蛋白質 / 時計タンパク質 / 小角散乱 (SAS) / サイズ排除クロマトグラフィ-X線小角散乱 / 超遠心分析(AUC)
Outline of Research at the Start

生物時計タンパク質として最もシンプル系であるシアノバクテリアのKaiA, KaiB, KaiCは、24時間周期で3者間での会合-解離を繰り返す。この挙動は6量体構造をもつKaiCの局所的な状態変化(ATP加水分解活性やリン酸化)と相関があるが、この「局所的な状態変化」と「3者間での会合-解離」を直接結ぶ分子間相互作用のメカニズムは解明されていない。本研究では、サイズ排除クロマトグラフィ-X線小角散乱測定による立体構造解析と、超遠心分析による構造安定性評価により、上記の2つを結びつけるメカニズムの解明を目指す。

Outline of Final Research Achievements

In this study, I investigated the structure of the clock protein “KaiC” in solution, which plays a fundamental role in circadian clock system of a cyanobacteria. First, an integral analytical method of analytical ultracentrifugation (AUC) and small-angle X-ray scattering (SAXS), namely “AUC-SAXS”, was developed to obtain the correct scattering profile for KaiC. Subsequently, the three-dimensional structure of KaiC, which reproduces the scattering profile, were searched with coarse grained-molecular dynamics (CG-MD). As a result, it was found that terminal chains of KaiC behaved as intrinsically disordered region.

Academic Significance and Societal Importance of the Research Achievements

本研究ではKaiCのKaiAおよびKaiBとの相互作用を理解するために重要な3次元構造の知見が得られた。今後は天然変性領域として振る舞う両末端鎖の運動に立脚した複合体解析によって、生物時計機構のより詳細な理解につながると期待される。
また、本研究で開発したAUC-SAXS法はKaiCだけに限らず一般的な生体高分子に対して普遍的に適用可能である。溶液中の凝集による影響はX線小角散乱実験において長年の問題であったため、これを解決する方法として幅広く応用されることが期待される。

Report

(3 results)
  • 2020 Annual Research Report   Final Research Report ( PDF )
  • 2019 Research-status Report
  • Research Products

    (26 results)

All 2021 2020 2019 Other

All Int'l Joint Research (2 results) Journal Article (7 results) (of which Int'l Joint Research: 2 results,  Peer Reviewed: 7 results,  Open Access: 7 results) Presentation (15 results) (of which Int'l Joint Research: 5 results) Book (1 results) Remarks (1 results)

  • [Int'l Joint Research] ANSTO(オーストラリア)

    • Related Report
      2020 Annual Research Report
  • [Int'l Joint Research] ILL(フランス)

    • Related Report
      2020 Annual Research Report
  • [Journal Article] Elucidation of the mechanism of subunit exchange in αB crystallin oligomers2021

    • Author(s)
      Inoue Rintaro、Sakamaki Yusuke、Takata Takumi、Wood Kathleen、Morishima Ken、Sato Nobuhiro、Okuda Aya、Shimizu Masahiro、Urade Reiko、Fujii Noriko、Sugiyama Masaaki
    • Journal Title

      Scientific Reports

      Volume: 11 Issue: 1 Pages: 1-9

    • DOI

      10.1038/s41598-021-82250-z

    • NAID

      120006956390

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Deuteration Aiming for Neutron Scattering2021

    • Author(s)
      Okuda Aya、Inoue Rintaro、Morishima Ken、Saio Tomohide、Yunoki Yasuhiro、Yagi-Utsumi Maho、Yagi Hirokazu、Shimizu Masahiro、Sato Nobuhiro、Urade Reiko、Kato Koichi、Sugiyama Masaaki
    • Journal Title

      Biophysics and Physicobiology

      Volume: 18 Issue: 0 Pages: 16-27

    • DOI

      10.2142/biophysico.bppb-v18.003

    • NAID

      130008000165

    • ISSN
      2189-4779
    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Solution structure of multi-domain protein ER-60 studied by aggregation-free SAXS and coarse-grained-MD simulation2021

    • Author(s)
      Okuda Aya、Shimizu Masahiro、Morishima Ken、Inoue Rintaro、Sato Nobuhiro、Urade Reiko、Sugiyama Masaaki
    • Journal Title

      Scientific Reports

      Volume: 11 Issue: 1 Pages: 1-13

    • DOI

      10.1038/s41598-021-85219-0

    • NAID

      120006997066

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] A feasibility study of inverse contrast-matching small-angle neutron scattering method combined with size exclusion chromatography using antibody interactions as model systems2021

    • Author(s)
      Sato Nobuhiro、Yogo Rina、Yanaka Saeko、Martel Anne、Porcar Lionel、Morishima Ken、Inoue Rintaro、Tominaga Taiki、Arimori Takao、Takagi Junichi、Sugiyama Masaaki、Kato Koichi
    • Journal Title

      The Journal of Biochemistry

      Volume: 0 Issue: 6 Pages: 1-8

    • DOI

      10.1093/jb/mvab012

    • NAID

      40022687772

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Integral approach to biomacromolecular structure by analytical-ultracentrifugation and small-angle scattering2020

    • Author(s)
      Morishima Ken、Okuda Aya、Inoue Rintaro、Sato Nobuhiro、Miyamoto Yosuke、Urade Reiko、Yagi-Utsumi Maho、Kato Koichi、Hirano Rina、Kujirai Tomoya、Kurumizaka Hitoshi、Sugiyama Masaaki
    • Journal Title

      Communications Biology

      Volume: 3 Issue: 1 Pages: 294-294

    • DOI

      10.1038/s42003-020-1011-4

    • NAID

      120006850271

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Dynamics of proteins with different molecular structures under solution condition2020

    • Author(s)
      Inoue Rintaro、Oda Takashi、Nakagawa Hiroshi、Tominaga Taiki、Saio Tomohide、Kawakita Yukinobu、Shimizu Masahiro、Okuda Aya、Morishima Ken、Sato Nobuhiro、Urade Reiko、Sato Mamoru、Sugiyama Masaaki
    • Journal Title

      Scientific Reports

      Volume: 10 Issue: 1 Pages: 21678-21678

    • DOI

      10.1038/s41598-020-78311-4

    • NAID

      120007193758

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Newly developed Laboratory-based Size exclusion chromatography Small-angle x-ray scattering System (La-SSS)2019

    • Author(s)
      R. Inoue, T. Nakagawa, K. Morishima, N. Sato, A. Okuda, R. Urade, R. Yogo, S. Yanaka, M. Yagi-Utsumi, K. Kato, K. Omoto, K. Ito, M. Sugiyama
    • Journal Title

      Scientific Reports

      Volume: 9 Issue: 1 Pages: 1-12

    • DOI

      10.1038/s41598-019-48911-w

    • NAID

      120006799252

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Open Access
  • [Presentation] 超遠心分析と小角散乱の統合手法(AUC-SAS)によるタンパク質複合体の構造解析2021

    • Author(s)
      守島健, 宮本洋祐, 清水将裕, 柚木康弘, 奥田綾, 佐藤信浩, 井上倫太郎, 裏出令子, 矢木宏和, 加藤晃一, 杉山正明
    • Organizer
      2020年度量子ビームサイエンスフェスタ
    • Related Report
      2020 Annual Research Report
  • [Presentation] ヒストンバリアントH2A.Bを含むヌクレオソームの新規動態の解析2021

    • Author(s)
      平野里奈, 有村泰宏, 鯨井智也, 柴田幹夫, 奥田綾, 守島健, 井上倫太郎, 杉山正明, 胡桃坂仁志
    • Organizer
      第38回染色体ワークショップ・第19回核ダイナミクス研究会
    • Related Report
      2020 Annual Research Report
  • [Presentation] Oligomeric structure and its temperature-dependent change of HspB1 from Chinese hamster ovary cell2020

    • Author(s)
      M. Yohda, M. Nakamura, N. Kurokawa, K. Morishima, R. Inoue, M. Sugiyama
    • Organizer
      2020 World Conference of Protein Science
    • Related Report
      2020 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Structural and functional characterization of sHsps from the fission yeast2020

    • Author(s)
      R. Nishijima, K. Morishima, R. Inoue, M. Sugiyama, M. Yohda
    • Organizer
      2020 World Conference of Protein Science
    • Related Report
      2020 Annual Research Report
    • Int'l Joint Research
  • [Presentation] 中性子散乱解析を目指したタンパク質ライゲーションの技術戦略2020

    • Author(s)
      奥田綾, 清水将裕, 守島健, 佐藤信浩, 井上倫太郎, 裏出令子, 杉山正明
    • Organizer
      日本中性子科学第20年会
    • Related Report
      2020 Annual Research Report
  • [Presentation] Innovative Small-Angle Scattering for Structural Analysis of Biomacromolecule in Solution2020

    • Author(s)
      守島健, 井上倫太郎, 中川達央, 宮本洋祐, 清水将裕, 奥田綾, 佐藤信浩, 裏出令子, 杉山正明
    • Organizer
      CBI2020大会
    • Related Report
      2020 Annual Research Report
  • [Presentation] タンパク質会合体の精密構造解析のための分析超遠心, SEC-SAXSおよびSEC-iCM-SANSの統合的利用2020

    • Author(s)
      佐藤信浩, 守島健, 井上倫太郎, 與語理那, 谷中冴子, 矢木宏和, 加藤晃一, Martel Anne, Porcar Lionel, 杉山正明
    • Organizer
      日本中性子科学第20年会
    • Related Report
      2020 Annual Research Report
  • [Presentation] X線小角散乱でみる大豆タンパク質のナノ構造に対する緩衝剤および塩の影響2020

    • Author(s)
      佐藤信浩, 裏出令子, 奥田綾, 守島健, 井上倫太郎, 杉山正明
    • Organizer
      日本農芸化学会2020年度大会
    • Related Report
      2020 Annual Research Report
  • [Presentation] 超遠心分析-小角散乱 (AUC-SAS) 法を用いた解離会合平衡下にあるタンパク質複合体の構造解析2020

    • Author(s)
      守島健, 矢木真穂, 井上倫太郎, 佐藤信浩, 奥田綾, 裏出令子, 加藤晃一, 杉山正明
    • Organizer
      2019年度量子ビームサイエンスフェスタ
    • Related Report
      2019 Research-status Report
  • [Presentation] 超遠心分析-小角散乱 (AUC-SAS) 法の高濃度凝集含有溶液への適用法の考察2020

    • Author(s)
      宮本洋祐, 守島健, 井上倫太郎, 佐藤信浩, 奥田綾, 裏出令子, 杉山正明
    • Organizer
      2019年度量子ビームサイエンスフェスタ
    • Related Report
      2019 Research-status Report
  • [Presentation] Structural characterization of the circadian clock protein complexes composed of KaiA, KaiB and KaiC by integrative structural approaches2019

    • Author(s)
      H. Yagi, Y. Yunoki, K. Morishima, K. Ishii, R. Murakami, L. Porcar, A. Martel, R. Inoue, K. Terauchi, S. Uchiyama, M. Sugiyama, K. Kato
    • Organizer
      J-PARC symposium
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] Subunit dynamics in a-crystallin through deuteration-assisted small-angle neutron scattering2019

    • Author(s)
      R. Inoue, Y. Sakamaki, K. Morishima, K. Wood, N. Sato, A. Okuda, R. Urade, N. Fujii, M. Sugiyama
    • Organizer
      J-PARC symposium
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] Creation and analysis of isomers-containing lens αA-crystallin2019

    • Author(s)
      T. Takata, K. Morishima, R. Inoue, M. Sugiyama, N. Fujii,
    • Organizer
      International Conference of Lens 2019
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] X線小角散乱を用いた大豆タンパク質の凝固に伴うナノ構造解析2019

    • Author(s)
      佐藤信浩, 裏出令子, 奥田綾, 守島健, 井上倫太郎, 杉山正明
    • Organizer
      日本食品科学工学会第66回大会
    • Related Report
      2019 Research-status Report
  • [Presentation] X線小角散乱と超遠心分析を協奏的に用いた蛋白質溶液の構造解析2019

    • Author(s)
      守島健, 井上倫太郎, 佐藤信浩, 裏出令子, 杉山正明
    • Organizer
      第19回日本蛋白質科学会年会
    • Related Report
      2019 Research-status Report
  • [Book] 62. 超遠心分析:相分離生物学の全貌(現代化学増刊46) 白木賢太郎編2020

    • Author(s)
      守島健
    • Total Pages
      400
    • Publisher
      東京化学同人
    • ISBN
      9784807913466
    • Related Report
      2020 Annual Research Report
  • [Remarks] 研究室ホームページ

    • URL

      https://www.rri.kyoto-u.ac.jp/PSlab/index.html

    • Related Report
      2019 Research-status Report

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Published: 2019-04-18   Modified: 2022-01-27  

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