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Tracking of protein folding transition path by single-molecule fluorescence measurements

Research Project

Project/Area Number 17K17608
Research Category

Grant-in-Aid for Young Scientists (B)

Allocation TypeMulti-year Fund
Research Field Physical chemistry
Biophysics
Research InstitutionTohoku University

Principal Investigator

Oikawa Hiroyuki  東北大学, 多元物質科学研究所, 助教 (40536778)

Research Collaborator Takahashi Satoshi  
Project Period (FY) 2017-04-01 – 2019-03-31
Project Status Completed (Fiscal Year 2018)
Budget Amount *help
¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000)
Fiscal Year 2018: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2017: ¥2,990,000 (Direct Cost: ¥2,300,000、Indirect Cost: ¥690,000)
Keywordsタンパク質 / フォールディング / 遷移経路 / 一分子蛍光測定 / 生物物理
Outline of Final Research Achievements

We aimed to track transition paths on protein folding events by single-molecule fluorescence resonance energy transfer (FRET) measurements. The time resolution of our existing line-confocal microscope combined with microfluidic chip was insufficient for tracking the transition events. To improve the time resolution, we built a new system based on hybrid photo detectors (HPD). By introducing HPD to the line-confocal microscope, we could obtain the single-molecule FRET traces with the time resolution of ten microsecond and the observation time of more than ten millisecond. As the target for the tracking of the transition path by using the new system, we tried to observe the fast conformational changes of F1-ATPase induced by the ATP hydrolysis. Unfortunately, although the conformational changes of F1-ATPase could not be observed, the method of single-molecule fluorescence measurement for tracking transition paths of biopolymer conformational changes was established.

Academic Significance and Societal Importance of the Research Achievements

タンパク質などの生体高分子は機能発現時に多くの場合立体構造が変化し、構造変化時にたどる遷移経路は酵素反応や分子生物学的過程と深く関わる。遷移経路の理解は、タンパク質が関わる疾病の治療や、人工タンパク質の創製などに有益であり、社会的に重要である。近年、分子動力学計算によって、計算機では生体分子の構造転移を追跡できるようになったが、実験的手法はほとんどない。本研究で開発した一分子の蛍光共鳴エネルギー移動(FRET)効率を10マイクロ秒分解能で追跡する手法はその有力な候補である。この手法は広範な生体高分子の構造変化の遷移経路追跡に適用可能であるため、学術的にも重要である。

Report

(3 results)
  • 2018 Annual Research Report   Final Research Report ( PDF )
  • 2017 Research-status Report
  • Research Products

    (12 results)

All 2018 2017

All Journal Article (3 results) (of which Peer Reviewed: 3 results,  Open Access: 1 results) Presentation (9 results) (of which Int'l Joint Research: 4 results,  Invited: 1 results)

  • [Journal Article] Microsecond resolved single-molecule FRET time series measurements based on the line confocal optical system combined with hybrid photodetectors2018

    • Author(s)
      Hiroyuki Oikawa、Takumi Takahashi、Supawich Kamonprasertsukac、Satoshi Takahashi
    • Journal Title

      Physical Chemistry Chemical Physics

      Volume: 20 Issue: 5 Pages: 3277-3285

    • DOI

      10.1039/c7cp06268k

    • Related Report
      2017 Research-status Report
    • Peer Reviewed
  • [Journal Article] Hypothesis: structural heterogeneity of the unfolded proteins originating from the coupling of the local clusters and the long-range distance distribution2018

    • Author(s)
      Takahashi Satoshi、Yoshida Aya、Oikawa Hiroyuki
    • Journal Title

      Biophysical Reviews

      Volume: 印刷中 Issue: 2 Pages: 363-373

    • DOI

      10.1007/s12551-018-0405-8

    • Related Report
      2017 Research-status Report
    • Peer Reviewed / Open Access
  • [Journal Article] Highly Heterogeneous Nature of the Native and Unfolded States of the B Domain of Protein A Revealed by Two-Dimensional Fluorescence Lifetime Correlation Spectroscopy2017

    • Author(s)
      Otosu Takuhiro、Ishii Kunihiko、Oikawa Hiroyuki、Arai Munehito、Takahashi Satoshi、Tahara Tahei
    • Journal Title

      The Journal of Physical Chemistry B

      Volume: 121 Issue: 22 Pages: 5463-5473

    • DOI

      10.1021/acs.jpcb.7b00546

    • Related Report
      2017 Research-status Report
    • Peer Reviewed
  • [Presentation] マイクロ秒分解一分子蛍光測定によるF1-ATPaseの構造変化追跡2018

    • Author(s)
      高橋巧、小井川浩之、須河光弘、高橋聡
    • Organizer
      第18回日本蛋白質科学会年会
    • Related Report
      2018 Annual Research Report
  • [Presentation] 単一分子蛍光分光法のための新しいタンパク質二重蛍光色素標識方法2018

    • Author(s)
      吉田文、金沢省、高橋泰人、 松井敏高、小井川浩之、浜田大三、高橋聡
    • Organizer
      第18回日本蛋白質科学会年会
    • Related Report
      2018 Annual Research Report
  • [Presentation] Single molecule fluorescence tracking at 10-μs resolution: Application to protein folding and functional dynamics2018

    • Author(s)
      Satoshi Takahashi、Hiroyuki Oikawa
    • Organizer
      第56回日本生物物理学会年会
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research / Invited
  • [Presentation] Ultra-fast dynamics of simple polyalanine peptides by using nanosecond region fluorescence correlation spectroscopy2018

    • Author(s)
      Supawich Kamonprasertsuk、Hiroyuki Oikawa、Satoshi Takahashi
    • Organizer
      第56回日本生物物理学会年会
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Investigation on the structural properties of proteins included in non-membraneous granule droplets2018

    • Author(s)
      Saya Nakano、Hiroyuki Oikawa、Satoshi Takahashi
    • Organizer
      第56回日本生物物理学会年会
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] Improvement of the fluorescent detected phage sorter for the application to phage display2018

    • Author(s)
      Hitomi Urabe、Saya Nakano、Yuki Shimizu、Naoki Mikosiba、Hiroyuki Oikawa、Satoshi Takahashi
    • Organizer
      第56回日本生物物理学会年会
    • Related Report
      2018 Annual Research Report
    • Int'l Joint Research
  • [Presentation] タンパク質構造変化の遷移経路追跡を目指した一分子蛍光測定装置の開発2017

    • Author(s)
      小井川 浩之、高橋 巧、須河 光弘、高橋 聡
    • Organizer
      第11回分子科学討論会
    • Related Report
      2017 Research-status Report
  • [Presentation] Development of Fluorescence Correlation Spectrometer for the Elucidation of Protein Interactions2017

    • Author(s)
      Asami Izaki、Hiroyuki Oikawa、Takeshi Tomita、Satoshi Takahashi
    • Organizer
      第55回日本生物物理学会年会
    • Related Report
      2017 Research-status Report
  • [Presentation] Unexpected heterogeneity and slow dynamics of simple poly-alanine peptides detected by single molecule fluorescence spectroscopy2017

    • Author(s)
      Supawich Kamonprasertsuk、Hiroyuki Oikawa、Satoshi Takahashi
    • Organizer
      第55回日本生物物理学会年会
    • Related Report
      2017 Research-status Report

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Published: 2017-04-28   Modified: 2020-03-30  

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