1989 Fiscal Year Annual Research Report

滑り運動系の微細構造

Research Project

Project/Area Number	01657004
Research Institution	Tokyo Metropolitan Organization for Medical Research
Principal Investigator	月田承一郎東京都臨床医学総合研究所, 超微形態, 研究員(室長) (50155347)
Co-Investigator(Kenkyū-buntansha)	若林克三大阪大学, 基礎工学部, 助手 (00029521) 盛田フミ北海道大学, 理学部, 教授 (80000818) 八木直人東北大学, 医学部, 助手 (80133940) 沼田治筑波大学, 生物科学系, 講師 (50189354) 岡本洋順天堂大学, 医学部, 講師 (70103877)
Keywords	骨格筋 / 筋収縮 / アクチン / ミオシン / ATP / エネルギ-転換 / 滑り運動 / 微細構造
Research Abstract	まずアクチン分子に関しては、沼田らによりテトラヒメナから単離されたアクチンのcDNAを酵母で発現させる系が初めて確立された。次にミオシン分子に関しては、山本の架橋実験により、アクチンフィラメントとの結合ドメイン(Kーsite)の解析が、盛田らの合成ポリペプチドを用いた方法により、Sーsiteと呼ばれるアクチン結合領域の解析がなされた。また岡本により、光アフィニティ-ラベル法を用いたミオシン分子のATPase活性中心の解析も精力的に行われた。滑り運動の構造的基盤を理解するには、上述したような、タンパク質分子そのものの詳しい構造を解析する一方で、滑り運動中の動的な構造を解析していく必要がある。そのための手法としては、電子顕微鏡法とX線回折法が現在のところ有用であると思われるが、そこで問題となるのはそれぞれの時間分解能である。まず、電子顕微鏡の時間分解能をさらに高めるために、caged化合物を用いた急速凍結法が月田らにより開発が試みられ、本年度中にほぼ完成した。このシステムを用いることにより、来年度から滑り運動系の動的微細構造に関してきわめて重要な情報が得られるものと期待される。次に、X線回折法は、より高い時間分解能と空間分解能を求めて、筑波高エネルギ-物理学研究所のシンクロトロン放射光を用いて詳細な実験が行われた。若林らは、イメ-ジングプレ-トをこの分野に初めて応用し、等尺性収縮中のミオシン分子とアクチン分子の動態についてきわめて明瞭な情報を得つつある。さらに八木らは、同様の方法により、短縮中の骨格筋のX線回折像を得ることに挑戦した。赤道反射に関してかなりの情報が集まった。今後、これら分担者が有機的につながり、共同研究を進めることで、当初の研究目的を充分達成できるものと考えている。

Research Products

(18 results)

All Other

All Publications (18 results)

[Publications] Tsukita,Sh.,Tsukita,Sa.: "Isolation of cellーtoーcell adherens junction from rat liver." J.Cell Biol.108. 31-41 (1989)
[Publications] Tsukita,Sa.,Hieda,Y.,Tsukita,Sh.: "A new 82ーkD barbed endーcapping protein(radixin)localized in the cellーtoーcell adherens junction: Purification and characterization" J.Cell Biol.108. 2369-2382 (1989)
[Publications] Hieda,Y.,Tsukita,Sa.,Tsukita,Sh.: "A new high molecular mass protein showing unique localization in desmosomal plaque." J.Cell Biol.109. 1511-1518 (1989)
[Publications] Tsukita,Sh.,Itoh,M.,Tsukita,Sa.: "A new 400 kDーprptein from isolated adherens junctions:Its localozation at the undercoat of adherens junctions and at microfilament bundles such as stress fibers and circumferential bundles." J.Cell Biol.109. 2905-2915 (1989)
[Publications] Matsumoto,G.,Tsukita,Sh.,Arai,T.: "Organization of the axonal cytoskeleton:Differentiation of the microtubule and actin filament arrays." In Cell Movement.Alan R.Liss Inc.335-356 (1989)
[Publications] Tsukita,Sh.,Tsukita,Sa.,Nagafuchi,A.: "The undercoat of adherens junctions:A key specialized structure in organogenesisi and carcinogenesis." Cell Struct.Funct.
[Publications] Matsubara,I.,Maughan,D.W.,Saeki,Y.,Yagi.N.: "Crossーbridge movement in rat cardiac muscle as a function of calcium concentration." J.Physiol.417. 555-565 (1989)
[Publications] Takahashi,M.,Fukushima,Y.,Inoue,K.,Hasegawa,Y.,Morita,F.,Takahashi,K.: "Ca^<++>ーsensitive transition in the molecular conformation of molluscan muscle myosins." J.Biochem.105. 149-151 (1989)
[Publications] Takahashi,M.,Morita,F.: "Myosin may stay in EADP species during the catch contraction in scallop smooth muscle." J.Biochem.106. 868-871 (1989)
[Publications] Suzuki,R.,Morita,F.,Nishi,N.,Tokura,S.: "Inhibition of actoーmyosine subfragmentー1 ATPase activity of analog peptides of the actinーbinding site around the Cys(SHI)of myosin heavy chain." J.Biol.Chem.(1990)
[Publications] Tregear,R.T.,Wakabayashi,K.,Tanaka,H.,Iwamoto,H.,Reedy,M.K.: "Xーray diffraction and electron microscopy from Lethocerus flight muscle partially relaxed by AMPPNP and ethylene glycol." J.Mal.Biol.(1990)
[Publications] Wakabayashi,K.,Amemiya,Y.: "Progress in Xーray synchrotron diffraction studies of muscle confraction." In Synchrotron Radiation Handbook(eds.S.Ebashi,E.Rubenstein,M.H.J.Koch). 4. (1990)
[Publications] Okamoto,Y.: "Localization of the esential light chain site of smooth muscle myosin after photoaffinity labeling." J.Musc,Res.Coll Motil.10. 258 (1989)
[Publications] Okamoto,Y.: "Contribution of the essential light chain of smooth muscle myosin for the active site." Prog.Clin.Biol.Res.315. 453-454 (1989)
[Publications] Yamamoto,K.: "Binding manner of actin to the lysineーrich sequence of myosin subfragmentー1 in the presence and absence of ATP" Biochemistry. 28. 5573-5577 (1989)
[Publications] Yamamoto.K.: "ATPーinduced structural change in myosin subfragmentー1 revealed by the location of protease cleavage sites on the primary structure." J.Mol.Biol.209. 703-709 (1989)
[Publications] Yamamoto,K.: "Shift of binding site at the interface between actin and myosin." Biochemistry. (1990)
[Publications] Yagi,N.,Matsubara,I.: "Structural changes in the thin filament during activation studies by Xーray diffraction of highly stretched skeletal muscle." J.Mol.Biol.208. 359-363 (1989)

1989 Fiscal Year Annual Research Report

滑り運動系の微細構造

Principal Investigator

月田 承一郎 東京都臨床医学総合研究所, 超微形態, 研究員(室長) (50155347)

Research Products

[Publications] Tsukita,Sh.,Tsukita,Sa.: "Isolation of cellーtoーcell adherens junction from rat liver." J.Cell Biol.108. 31-41 (1989)

[Publications] Tsukita,Sa.,Hieda,Y.,Tsukita,Sh.: "A new 82ーkD barbed endーcapping protein(radixin)localized in the cellーtoーcell adherens junction: Purification and characterization" J.Cell Biol.108. 2369-2382 (1989)

[Publications] Hieda,Y.,Tsukita,Sa.,Tsukita,Sh.: "A new high molecular mass protein showing unique localization in desmosomal plaque." J.Cell Biol.109. 1511-1518 (1989)

[Publications] Tsukita,Sh.,Itoh,M.,Tsukita,Sa.: "A new 400 kDーprptein from isolated adherens junctions:Its localozation at the undercoat of adherens junctions and at microfilament bundles such as stress fibers and circumferential bundles." J.Cell Biol.109. 2905-2915 (1989)

[Publications] Matsumoto,G.,Tsukita,Sh.,Arai,T.: "Organization of the axonal cytoskeleton:Differentiation of the microtubule and actin filament arrays." In Cell Movement.Alan R.Liss Inc.335-356 (1989)

[Publications] Tsukita,Sh.,Tsukita,Sa.,Nagafuchi,A.: "The undercoat of adherens junctions:A key specialized structure in organogenesisi and carcinogenesis." Cell Struct.Funct.

[Publications] Matsubara,I.,Maughan,D.W.,Saeki,Y.,Yagi.N.: "Crossーbridge movement in rat cardiac muscle as a function of calcium concentration." J.Physiol.417. 555-565 (1989)

[Publications] Takahashi,M.,Fukushima,Y.,Inoue,K.,Hasegawa,Y.,Morita,F.,Takahashi,K.: "Ca^<++>ーsensitive transition in the molecular conformation of molluscan muscle myosins." J.Biochem.105. 149-151 (1989)

[Publications] Takahashi,M.,Morita,F.: "Myosin may stay in EADP species during the catch contraction in scallop smooth muscle." J.Biochem.106. 868-871 (1989)

[Publications] Suzuki,R.,Morita,F.,Nishi,N.,Tokura,S.: "Inhibition of actoーmyosine subfragmentー1 ATPase activity of analog peptides of the actinーbinding site around the Cys(SHI)of myosin heavy chain." J.Biol.Chem.(1990)

[Publications] Tregear,R.T.,Wakabayashi,K.,Tanaka,H.,Iwamoto,H.,Reedy,M.K.: "Xーray diffraction and electron microscopy from Lethocerus flight muscle partially relaxed by AMPPNP and ethylene glycol." J.Mal.Biol.(1990)

[Publications] Wakabayashi,K.,Amemiya,Y.: "Progress in Xーray synchrotron diffraction studies of muscle confraction." In Synchrotron Radiation Handbook(eds.S.Ebashi,E.Rubenstein,M.H.J.Koch). 4. (1990)

[Publications] Okamoto,Y.: "Localization of the esential light chain site of smooth muscle myosin after photoaffinity labeling." J.Musc,Res.Coll Motil.10. 258 (1989)

[Publications] Okamoto,Y.: "Contribution of the essential light chain of smooth muscle myosin for the active site." Prog.Clin.Biol.Res.315. 453-454 (1989)

[Publications] Yamamoto,K.: "Binding manner of actin to the lysineーrich sequence of myosin subfragmentー1 in the presence and absence of ATP" Biochemistry. 28. 5573-5577 (1989)

[Publications] Yamamoto.K.: "ATPーinduced structural change in myosin subfragmentー1 revealed by the location of protease cleavage sites on the primary structure." J.Mol.Biol.209. 703-709 (1989)

[Publications] Yamamoto,K.: "Shift of binding site at the interface between actin and myosin." Biochemistry. (1990)

[Publications] Yagi,N.,Matsubara,I.: "Structural changes in the thin filament during activation studies by Xーray diffraction of highly stretched skeletal muscle." J.Mol.Biol.208. 359-363 (1989)

月田承一郎東京都臨床医学総合研究所, 超微形態, 研究員(室長) (50155347)