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2008 Fiscal Year Final Research Report

Development of an NMR strategy for the structural analysis of themolecular recognition in the membrane protein complex in lipidbilayers

Research Project

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Project/Area Number 19590037
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Physical pharmacy
Research InstitutionThe University of Tokyo

Principal Investigator

OSAWA Masanori  The University of Tokyo, 大学院・薬学系研究科, 助教 (60361606)

Co-Investigator(Kenkyū-buntansha) SHIMADA Ichio  東京大学, 大学院・薬学系研究科, 教授 (70196476)
Project Period (FY) 2007 – 2008
Keywords膜蛋白質 / 分子認識 / NMR / 構造生物学
Research Abstract

脂質二重膜中の膜蛋白質はビーズに固定化することにより、安定性が劇的に向上するが、不溶性成分の共存によりNMRシグナルの広幅化・感度の低下が問題となっていた。そこで、マジック角回転(MAS)による高感度化・高分解能化を目指した。
本研究では、高速MASに耐えることができ、広い固・液界面が確保可能な固定化担体を幅広く比較検討した結果、CM-シリカビーズを見出した。
次に、ユビキチン(Ub)と酵母Ub水酸化酵素(YUH)の相互作用系において、YUHをCMシリカビーズに固定化し、不均一系超分子を模倣した系を作成した。このYUH固定化Zn-NTAシリカビーズを均一2H15N標識したUbの溶液に懸濁し、MAS 条件下で転移交差飽和法の測定を試みたところ、従来法では解析が困難であった固体と液体が共存した超分子系において、Ub 上のYUH 結合界面残基の同定に成功した。

  • Research Products

    (21 results)

All 2008 2007

All Journal Article (13 results) (of which Peer Reviewed: 12 results) Presentation (4 results) Book (4 results)

  • [Journal Article] Stable isotope labeling of proteinby Kluyveromyces lactis for NMR study2008

    • Author(s)
      Sugiki T., Shimada I., Takahashi H.
    • Journal Title

      J Biomol NMR 42

      Pages: 159-162

    • Peer Reviewed
  • [Journal Article] Carbohydrate bindingmechanism of the macrophagegalactose-type C-type lectin 1 revealedby saturation transfer experiments2008

    • Author(s)
      Sakakura M., Oo-Puthinan S., MoriyamaC., Kimura T., Moriya J., Irimura T., Shimada I.
    • Journal Title

      J Biol Chem 283

      Pages: 33665-33673

    • Peer Reviewed
  • [Journal Article] Chondroitin sulfate E fragmentsenhance CD44 cleavage andCD44-dependent motility in tumor cells2008

    • Author(s)
      Sugahara K. N., Hirata T., Tanaka T., Ogino S., Takeda M., Terasawa H., Shimada I, Tamura J, ten Dam GB, vanKuppevelt TH, Miyasaka M.
    • Journal Title

      Cancer Res. 68

      Pages: 7191-7199

    • Peer Reviewed
  • [Journal Article] Amino acidselective cross-saturation method foridentification of proximal residue pairsin a protein-protein complex2008

    • Author(s)
      Igarashi S., Osawa M., Takeuchi K., Ozawa S., Shimada I.
    • Journal Title

      J Am Chem Soc. 130

      Pages: 12168-12176

    • Peer Reviewed
  • [Journal Article] Structural insights into activation ofphosphatidylinositol 4-kinase (Pik1) byyeast frequenin (Frq1)2007

    • Author(s)
      Strahl T, Huttner IG, Lusin JD, OsawaM, King D, Thorner J, Ames JB.
    • Journal Title

      J Biol Chem. 282(42)

      Pages: 30949-59

    • Peer Reviewed
  • [Journal Article] Structuralbasis of the collagen-binding mode ofdiscoidin domain receptor 22007

    • Author(s)
      Ichikawa O, Osawa M, Nishida N,Goshima N, and Shimada I.
    • Journal Title

      EMBO J 26

      Pages: 4168-4176

  • [Journal Article] Mechanism of mRNA deadenylation : evidence for a molecular interplaybetween translation termination factoreRF3 and mRNA deadenylases2007

    • Author(s)
      Funakoshi Y, Doi Y, Hosoda N, Uchida N,Osawa M, Shimada I, Tsujimoto M,Suzuki T, Katada T, Hoshino S.
    • Journal Title

      Genes Dev. 21(23)

      Pages: 3135-48

    • Peer Reviewed
  • [Journal Article] Nickel binding to NikA : anadditional binding site reconcilesspectroscopy, calorimetry andcrystallography2007

    • Author(s)
      Addy C, Ohara M, Kawai F, Kidera A,Ikeguchi M, Fuchigami S, Osawa M,Shimada I, Park SY, Tame JR andHeddle JG.
    • Journal Title

      Acta Crystallogr D BiolCrystallogr 63

      Pages: 221-229

    • Peer Reviewed
  • [Journal Article] Bovin NV and Irimura T. The aminoacids involved in the distinctcarbohydrate specificities betweenmacrophage galactose-type C-typelectins 1 and 2 (CD301a and b) of mice2007

    • Author(s)
      Oo-puthinan S, Maenuma K, SakakuraM, Denda-Nagai K, Tsuiji M, Shimada I, Nakamura-Tsuruta S, and Hirabayashi S.
    • Journal Title

      Biochim Biophys Acta 1780(2)

      Pages: 89-100

    • Peer Reviewed
  • [Journal Article] Structural basisof the transition-state stabilization inantibody-catalyzed hydrolysis2007

    • Author(s)
      Sakakura M, Takahashi H, Shimba N,Fujii I, and Shimada I.
    • Journal Title

      J Mol Biol 367

      Pages: 133-147

    • Peer Reviewed
  • [Journal Article] PairwiseNMR Experiments for theDetermination of Protein BackboneDihedral Angle Based onCross-Correlated Spin Relaxation.2007

    • Author(s)
      Takahashi H and Shimada I.
    • Journal Title

      JBiomol NMR 37

      Pages: 179-185

    • Peer Reviewed
  • [Journal Article] Backboneresonance assignment for the outermembrane lipoprotein receptor LolBfrom Escherichia coli2007

    • Author(s)
      Nakada S, Sakakura M, Takahashi H,Tokuda H and Shimada I.
    • Journal Title

      BiomolecularNMR Assignments 1

      Pages: 121-123

    • Peer Reviewed
  • [Journal Article] Backboneresonance assignments for theperiplasmic chaperone LolA fromEscherichia coli2007

    • Author(s)
      Nakada S, Takahashi H, Sakakura M,Kurono M and Shimada I.
    • Journal Title

      Biomolecular NMRAssignments 1

      Pages: 125-127

    • Peer Reviewed
  • [Presentation] 生体超分子における分子認識機構解明に向けたHR-MASの適用2008

    • Author(s)
      豊永 翔
    • Organizer
      第47回NMR 討論会
    • Place of Presentation
      茨城県つくば市
    • Year and Date
      2008-11-13
  • [Presentation] 巨大タンパク質複合体のモデル構築を目的とした残基選択的交差飽和法の開発2008

    • Author(s)
      小澤新一郎
    • Organizer
      第47回NMR 討論会
    • Place of Presentation
      茨城県つくば市
    • Year and Date
      2008-11-13
  • [Presentation] Structural studies onthe interaction of spermine withinwardly rectifying potassium channel2008

    • Author(s)
      Osawa M. et.al.
    • Organizer
      XXIIIrd International Conference onMagnetic Resonance in BiologicalSystems
    • Place of Presentation
      San Diego, CA, USA
    • Year and Date
      2008-08-24
  • [Presentation] Structural basis of the gating of G protein-activatedinwardly rectifying potassium channelby Gbg2008

    • Author(s)
      Yokogawa M. et.al.
    • Organizer
      XXIIIrd InternationalConference on Magnetic Resonance in Biological Systems
    • Place of Presentation
      San Diego, CA, USA
    • Year and Date
      2008-08-24
  • [Book] NMRによるソフトな分子間相互作用解析法の開発2008

    • Author(s)
      松本昌彦、上田卓見、嶋田一夫
    • Total Pages
      959-971
    • Publisher
      生化学
  • [Book] コラーゲン結合タンパク質を介した生命プロセスの活性化機構2008

    • Author(s)
      西田紀貴、嶋田一夫
    • Total Pages
      483-492
    • Publisher
      生化学
  • [Book] 「等温滴定型カロリメトリー(ITC)-熱量測定による相互作用の定量的解析-」実験医学別冊「生命科学のための機器分析実験ハンドブック2007

    • Author(s)
      大澤匡範、嶋田一夫
    • Total Pages
      249-253
    • Publisher
      羊土社
  • [Book] 交換系利用NMRによるリガンド分子の標的分子結合状態の構造解析 高橋栄夫・嶋田一夫2007

    • Author(s)
      高橋栄夫、嶋田一夫
    • Total Pages
      959-965
    • Publisher
      蛋白質・核酸・酵素

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Published: 2010-06-10   Modified: 2016-04-21  

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