High-through put isolation of functional mutants of actin using Dictyostelium expression system

• Project/Area Number: 22570163
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Biophysics
• Research Institution: Teikyo University
• Principal Investigator: WAKABAYASHI Takeyuki 帝京大学, 医療技術学部, 教授 (90011717)
• Project Period (FY): 2010 – 2012
• Project Status: Completed (Fiscal Year 2012)
• Budget Amount: ¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000); Fiscal Year 2012: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2011: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2010: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
• Keywords: アクチン / ミオシン / 細胞分化 / 収縮タンパク質 / 機能的変異株 / 細胞性粘菌 / X 線結晶解析 / ATPase / アクチン重合 / アクチン・ミオシン相互作用 / X線結晶解析 / アクチン変異体 / ディクチオ型粘菌 / 胞子形成能 / アクチンフィラメント / アクチン重合能

Research Abstract

Actin filaments are found in all eukaryotic cells and are essentialfor many of their movements including cell migration and cell division. We found that the cell behavior of Dictyostelium cells changed, when the vector for the expression of mutant actin was incorporated. Using this phenomenon, functional mutants of actin were isolated efficiently. It was found that the tyrosine-143 of actin is important for actin assembly and the activation of myosin ATPase. The crystal structures of such mutants were determined.

Research Products

• [Journal Article] Electron microscopic evidence for the myosin head lever arm mechanism inhydrated myosin filaments using the gas environmental chambe (2011)
  • Author(s): H. Minoda, T. Okabe, Y. Inayoshi, T. Minakata, Y. Miyauchi, M. Tanokura, E. Katayama, T. Wakabayashi, T. Akimoto, *H. Sugi
  • Journal Title: Biochem. Biophys. Res. Commun Volume: 405 Pages: 651-656
  • Peer Reviewed
• [Journal Article] アクチンのフィラメント構造と重合機構---重合による ATPase 活性化のメカニズム--- (2011)
  • Author(s): 若林健之、村上健次
  • Journal Title: 生物物理 Volume: 51(6) Pages: 256-259
  • NAID: 10030037949
  • Peer Reviewed
• [Journal Article] Structural Basis for Actin Assembly (2010)
  • Author(s): K. Murakami, T. Yasunaga, T. Q. P. Noguchi, Y. Gomibuchi, K. X. Ngo, T. Q. P. Uyeda, & T. Wakabayashi
  • Journal Title: Activation of ATP Hydrolysis, and Delayed Phosphate Release Volume: 143 Pages: 275-287
  • Peer Reviewed
• [Journal Article] Dominant negative mutant actins identified in flightless Drosophila can be classified into three classes, including perturbation of the tropomyosin-troponin system (2010)
  • Author(s): T. Q. P. Noguchi, Y. Gomibuchi, K. Murakami, H. Ueno, K. Hirose, T. Wakabayashi, & T. Q. P. Uyeda
  • Journal Title: J. Biol. Chem Volume: 285 Pages: 4337-4347
  • Peer Reviewed
• [Journal Article] Structural Basis for Actin Assembly, Activation of ATP Hydrolysis, and Delayed Phosphate Release. (2010)
  • Author(s): K.Murakami, T.Yasunaga, T.Q.P.Noguchi, Y.Gomibuchi, K.X.Ngo, T.Q.P.Uyeda, T.Wakabayashi
  • Journal Title: Cell Volume: 143 Pages: 275-287
  • Peer Reviewed
• [Journal Article] Dominant negative mutant actins identified in flightless Drosophila can be classified into three classes, including perturbation of the tropomyosin-troponin system. (2010)
  • Author(s): T.Q.P.Noguchi, Y.Gomibuchi, K.Murakami, H.Ueno, K.Hirose, T.Wakabayashi, T.Q.P.Uyeda
  • Journal Title: J.Biol.Chem Volume: 285 Pages: 4337-4347
  • Peer Reviewed
• [Presentation] Changes of biochemical properties of Dictyostelium actin induced by mutation of tyrosine-143 (2012)
  • Author(s): 五味渕由貴、 上田太郎、 若林健之
  • Organizer: 日本生物物理学会
  • Place of Presentation: Nagoya
• [Presentation] Changes of biochemical properties of Dictyostelium actin induced by mutation of tyrosine-143 (2012)
  • Author(s): T. Wakabayashi and K. Murakami
  • Organizer: Interaction between the Components of Actin-containing Filaments
  • Place of Presentation: First Kyushu Annual Meeting of Biophysics Society of Japan
• [Presentation] Changes of biochemical properties of Dictyostelium actin induced by mutation of tyrosine-143 (2012)
  • Author(s): T. Wakabayashi
  • Organizer: 日本生物物理学会
  • Place of Presentation: 名古屋大学・東山キャンパス
• [Presentation] (2011)
  • Author(s): 若林健之、村上健次、五味渕由貴、安永卓生、上田太郎、野口太郎
  • Organizer: 蛋白質科学会年会ワークショップ
  • Place of Presentation: 大阪
  • Year and Date: 2011-06-07
• [Presentation] Structural Basis of Actin Polymerization, ATPase activation and Delayed Phosphate Release (2010)
  • Author(s): K. Murakami, T. Yasunaga, T. Q. P. Noguchi, Y. Gomibuchi, K. Ngo, T. Q. P. Uyeda, and T. Wakabayashi
  • Organizer: Oral Presentation The 48_<th> Biophysics Society of Japan Annual meeting
  • Place of Presentation: Sendai
  • Year and Date: 2010-09-20
• [Presentation] How actin assembly triggers activation of ATP hydrolysis (2010)
  • Author(s): K. Murakami, T. Yasunaga, T. Q. P. Noguchi, T. Q. P. Uyeda & T . Wakabayashi
  • Organizer: Symposium on motor functions
  • Place of Presentation: Alpbach, Austria