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Structural and functional analysis of the motor protein prestin using purified protein

Research Project

Project/Area Number 18390455
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field Otorhinolaryngology
Research InstitutionTohoku University

Principal Investigator

WADA Hiroshi  Tohoku University, Graduate School of Engineering, Professor (30111264)

Co-Investigator(Kenkyū-buntansha) KOBAYASHI Toshimitsu  Graduate School of Medicine, 大学院・医学系研究科, professor (80133958)
KUMAGAI Izumi  Graduate School of Engineering, 大学院・工学研究科, Professor (10161689)
IKEDA Katsuhisa  Department of Otorhinolaryngology, Juntendo University School of Medicine, Professor (70159614)
TSUMOTO Kouhei  Department of Medical Genome Sciences, The University of Tokyo, Associate Professor (90271866)
Project Period (FY) 2006 – 2007
Project Status Completed (Fiscal Year 2007)
Budget Amount *help
¥16,380,000 (Direct Cost: ¥14,100,000、Indirect Cost: ¥2,280,000)
Fiscal Year 2007: ¥9,880,000 (Direct Cost: ¥7,600,000、Indirect Cost: ¥2,280,000)
Fiscal Year 2006: ¥6,500,000 (Direct Cost: ¥6,500,000)
KeywordsOtology / Protein / Molecular motor / Prestin / Atomic force microscopy / Purification / Isothermal titration calorimetry / 生物物理 / 走査プローブ顕微鏡 / パッチクランプ法 / 等温滴定型カロリメトリー / 国際研究者交流 / アメリカ / 膜タンパク質
Research Abstract

Outer hair cells (OHCs), which are sensory cells in the mammalian cochlea, are able to elongate and contract in length in response to changes in memebrane potential. This motility is believed to amplify the motion of the cochlear partition generated by acoustical stimulation. As a result, mammalian hearing is characterized by high sensitivity, wide dynamic range and sharp frequency selectivity. The origin of this motility is associated with conformational change of the motor protein prestin expressed in the lateral wall of OHCs. In the present study, an attempt was made to elucidate the molecular structure and molecular mechanisms of the conformational changes of prestin. First, the plasma membranes of prestin-transfected Chinese hamster ovary (CHO) cells and those of untransfected CHO cells were observed by atomic force microscopy (AFM). More particle-like structures with a diameter of 8-18 nm were found to exist in the plasma membrane of the prestin-transfected CHO cells than in that of the untransfected CHO cells, indicating that the diameter of prestin is 8-12 nm. Second, prestin molecules expressed in the plasma membranes of prestin-transfected CHO cells were labeled with Quantum dots (Qdots), and those dots, about 8 nm in height, were clearly imaged by AFM. Squarish ring-like structures, each with four peaks and one valley at its center, were observed in the vicinity of the Qdots, suggesting that prestin forms a tetramer in the plasma membranes of the prestin-transfected CHO cells. Finally, prestin molecules were purified from prestin-transfected CHO cells. To reveal whether the purified prestin was functional, based on a finding that prestin binds anions, the interaction between purified prestin and chloride was examined by isothermal titration calorimetry. As a result, heat absorption resulting from prestin-chloride interaction was detected, suggesting that the obtained purified prestin was functional.

Report

(3 results)
  • 2007 Annual Research Report   Final Research Report Summary
  • 2006 Annual Research Report
  • Research Products

    (35 results)

All 2008 2007 2006 Other

All Journal Article (7 results) (of which Peer Reviewed: 3 results) Presentation (24 results) Book (3 results) Remarks (1 results)

  • [Journal Article] Effects of heat stress on filamentous actin and prestin of outer hair cells in mice2007

    • Author(s)
      Yoko Kitsunai
    • Journal Title

      Brain Research 1177

      Pages: 47-58

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Recent findings on our auditory system: it is highly sensitiveowing to the motility of sensory cells2007

    • Author(s)
      Hiroshi Wada
    • Journal Title

      International Journal of Acoustics and Vibration 12

      Pages: 75-82

    • Related Report
      2007 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Imaging by atomic force microscopy of the plasma membrane of prestin-transfected Chinese hamster ovary cells2006

    • Author(s)
      Michio Murakoshi
    • Journal Title

      Journal of the Association for Research in Otolaryngology 7

      Pages: 267-278

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary 2006 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Imaging by atomic force microscopy of the plasma membrane of prestin-transfected Chinese hamster ovary cells2006

    • Author(s)
      Michio Murakoshi et. al.
    • Journal Title

      Journal of the Association for Research in Otolaryngology 7

      Pages: 267-278

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Journal Article] Local mechanical properties of mouse outer hair cells : Atomic force microscopic study2006

    • Author(s)
      Michio Murakoshi
    • Journal Title

      Auris Nasus Larynx 33

      Pages: 149-157

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Effects of heat stress on Young's modulus of outer hair cells in mice2006

    • Author(s)
      Michio Murakoshi
    • Journal Title

      Brain Research 1107

      Pages: 121-130

    • Related Report
      2006 Annual Research Report
  • [Journal Article] Analysis of the dynamic behavior of the inner hair cell stereocilia by the finite element method2006

    • Author(s)
      Toshihiro Matsui
    • Journal Title

      JSME International Journal 49C

      Pages: 828-836

    • Related Report
      2006 Annual Research Report
  • [Presentation] Immune Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells Using Quantum Dots2008

    • Author(s)
      Michio Murakoshi
    • Organizer
      The 31st Association for Research in Otolaryngology Midwinter meeting
    • Place of Presentation
      Phoenix, U.S.A.
    • Year and Date
      2008-02-19
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Immune Atomic Force Microscopy of the Plasma Membrane of Prestin-Transfected Chinese Hamster Ovary Cells Using Quantum Dots2008

    • Author(s)
      Michio Murakoshi
    • Organizer
      The 31st Association for Research in Otolaryngology Midwinter meeting
    • Place of Presentation
      Phoenix, U.S.A
    • Year and Date
      2008-02-19
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] 量子ドットで標識したタンパク質モータプレスチンの原子間力顕微鏡による可視化2008

    • Author(s)
      村越 道生
    • Organizer
      日本機械学会第20回バイオエンジニアリング講演会
    • Place of Presentation
      東京
    • Year and Date
      2008-01-25
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Imaging by Atomic Force Microscopy of the Motor Protein Prestin Labeled with Quantum Dots2008

    • Author(s)
      Michio Murakoshi
    • Organizer
      The 20th Bioengineering Conference, 2006 Anual meeting of BED/JSME
    • Place of Presentation
      Tokyo
    • Year and Date
      2008-01-25
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Heterologous expression of the outer hair cell motor protein prestin in Sf9 cells2008

    • Author(s)
      Takashi Tadenuma
    • Organizer
      The 31st Association for Research in Otolaryngology Midwinter meeting
    • Place of Presentation
      Phoenix
    • Related Report
      2007 Annual Research Report
  • [Presentation] Enthalpy measurement of anion binding to the motor protein prestin byisothermal titration calorimetry.2007

    • Author(s)
      Koji Iida
    • Organizer
      The 3rd Tohoku-NUS Joint Symposium on Nano-Biomedical Engineering in the East Asian-Pacific Rim Region. Nano-Biomedical Engineering in the East Asian-Pacific Rim Region
    • Place of Presentation
      Singapore
    • Year and Date
      2007-12-11
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Enthalpy measurement of anion binding to the motor protein prestin byisothermal titration calorimetry2007

    • Author(s)
      Koji Iida
    • Organizer
      The 3rd Tohoku-NUS Joint Symposium on Nano-Biomedical Engineering in the East Asian-Pacific Rim Region. Nano- Biomedical Engineering in the East Asian-Pacific Rim Region
    • Place of Presentation
      Singapore
    • Year and Date
      2007-12-11
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Interaction of the inner ear motor protein prestin with chloride studiedby isothermal titration calorimetry2007

    • Author(s)
      Koji Iida
    • Organizer
      Third Asian Pacific Conference on Biomechanics
    • Place of Presentation
      Tokyo
    • Year and Date
      2007-11-07
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] タンパク質モータprestinの精製2007

    • Author(s)
      飯田 浩司
    • Organizer
      第17回日本耳科学会総会
    • Place of Presentation
      福岡
    • Year and Date
      2007-10-18
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Annual Research Report 2007 Final Research Report Summary
  • [Presentation] Purification of the motor protein prestin2007

    • Author(s)
      Koji Iida
    • Organizer
      Anual meeting of the Otological Society of Japan
    • Place of Presentation
      Hukuoka
    • Year and Date
      2007-10-18
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Isothermal titration calorimetry study of the interactions between purified prestin and chloride2007

    • Author(s)
      Koji Iida
    • Organizer
      6th Molecular Biology Hearing & Deafness Conference
    • Place of Presentation
      Hinxton, UK
    • Year and Date
      2007-07-11
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Isothermal titration calorimetry study of the interactions between purified prestin and chloride2007

    • Author(s)
      Koji Iida
    • Organizer
      6th Molecular Biology Hearing & Deafness Conference
    • Place of Presentation
      Hinton, UK
    • Year and Date
      2007-07-11
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Purification and isothermal titration calorimetric analysis of the motor protein prestin.2007

    • Author(s)
      Koji Iida
    • Organizer
      JCA2007 The Japan-China Joint Conference on Acoustics
    • Place of Presentation
      Sendai
    • Year and Date
      2007-06-05
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Annual Research Report 2007 Final Research Report Summary
  • [Presentation] Purification and isothermal titration calorimetric analysis of the motor protein prestin2007

    • Author(s)
      Koji Iida
    • Organizer
      JCA2007 The Japan-China Joint Conference on Acoustics
    • Place of Presentation
      Sendai
    • Year and Date
      2007-06-05
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Characterization of the interaction between the motor protein prestin and anions by isothermal titration calorimetry2007

    • Author(s)
      Koji Iida
    • Organizer
      The 9th International Symposium on Future Medical Engineering based on Bio-nanotechnology
    • Place of Presentation
      Sendai
    • Year and Date
      2007-01-09
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] タンパク質モータプレスチンと塩化物イオンの相互作用解析2007

    • Author(s)
      飯田 浩司
    • Organizer
      日本機械学会第19回バイオエンジニアリング講演会
    • Place of Presentation
      仙台
    • Year and Date
      2007-01-07
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Interaction Between the Motor Protein Prestin and Chloride2007

    • Author(s)
      Koji Iida
    • Organizer
      The 19th Bioengineering Conference, 2006 Anual meeting of BED/JSME
    • Place of Presentation
      Sendai
    • Year and Date
      2007-01-07
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Structure and function of human auditory system: its high sensitivity is achieved by cochlear amplification.2007

    • Author(s)
      Hiroshi Wada
    • Organizer
      The 3rd Tohoku-NUS Joint Symposium on Nano-Biomedical Engineering in the East Asian-Pacific Rim Region. Nano-Biomedical Engineering in the East Asian-Pacific Rim Region
    • Place of Presentation
      Singapore
    • Related Report
      2007 Annual Research Report
  • [Presentation] Enthalpy measurement of anion binding to the motor protein prestin by isothermal titration calorimetry.2007

    • Author(s)
      Koji Iida
    • Organizer
      The 3rd Tohoku-NUS Joint Symposium on Nano-Biomedical Engineering in the East Asian-Pacific Rim Region. Nano-Biomedical Engineering in the East Asian-Pacific Rim Region
    • Place of Presentation
      Singapore
    • Related Report
      2007 Annual Research Report
  • [Presentation] Structural analysis of the motor protein prestin.2007

    • Author(s)
      Hiroshi Wada
    • Organizer
      Symposium on Biomechanics at Micro-and Nanoscale Levels
    • Place of Presentation
      Tokyo
    • Related Report
      2007 Annual Research Report
  • [Presentation] Interaction of the inner ear motor protein prestin with chloride studied by isothermal titration calorimetry2007

    • Author(s)
      Koji Iida
    • Organizer
      Third Asian Pacific Conference on Biomechanics
    • Place of Presentation
      Tokyo
    • Related Report
      2007 Annual Research Report
  • [Presentation] Isothermal titration calorimetry study of the interactions between purified prestin and chloride2007

    • Author(s)
      Koji Iida
    • Organizer
      6th Molecular Biology Hearing & Deafness Conference
    • Place of Presentation
      Hinxton
    • Related Report
      2007 Annual Research Report
  • [Presentation] Purification of the motor protein prestin and its interaction with chloride studied by isothermal titration calorimetry2006

    • Author(s)
      Koji Iida
    • Organizer
      The 2nd Shanghai International Conference on Physiological Biophysics-Audition and Vision
    • Place of Presentation
      Shanghai, China
    • Year and Date
      2006-11-06
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Presentation] Purification of the motor protein prestin and its interaction with chloride studied by isothermal titration calorimetry2006

    • Author(s)
      Koji Iida
    • Organizer
      The 2nd Shanghai International Conference on Physiological Biophysics - Audition and Vision
    • Place of Presentation
      Shanghai, China
    • Year and Date
      2006-11-06
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Book] Molecular Protocols in Auditory Research2008

    • Author(s)
      Walker, J
    • Publisher
      Humana Press(In press)
    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Book] Molecular Protocols in Auditory Research2008

    • Author(s)
      Walker, J (ed.)
    • Publisher
      Humana Press
    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2007 Final Research Report Summary
  • [Book] Handbook of Noise and Vibration Control2007

    • Author(s)
      Croker, M. J.
    • Publisher
      John Wiley & Sons Inc
    • Related Report
      2007 Annual Research Report
  • [Remarks] 「研究成果報告書概要(和文)」より

    • URL

      http://www.wadalab.mech.tohoku.ac.jp/index.html

    • Related Report
      2007 Final Research Report Summary

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Published: 2006-04-01   Modified: 2016-04-21  

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