1990 Fiscal Year Annual Research Report

生体膜機能分子のリン脂質平面膜法による解析

Research Project

Project/Area Number	02304067
Research Institution	Jichi Medical University
Principal Investigator	平田肇自治医科大学, 医学部, 講師 (40049052)
Co-Investigator(Kenkyū-buntansha)	高木雅行大阪大学, 教養部, 助教授 (20028145) 曽我部正博名古屋大学, 医学部, 助教授 (10093428) 津田基之姫路工業大学, 理学部, 教授 (60045458) 桐野豊九州大学, 薬学部, 教授 (10012668) 葛西道生大阪大学, 基礎工学部, 教授 (40022595)
Keywords	リン脂質平面膜 / イオンチャンネル / イオンポンプ / 情報伝達 / 生体膜機能素子 / 合成ペプチド / クライオ電子顕微鏡 / コンピュ-タ-シミュレ-ション
Research Abstract	本研究は、イオンチャンネルやイオンポンプなど生体膜機能分子の活性部位や活性調節部位など分子内部の構造と機能の連関を探ることを目標に、リン脂質平面膜法を主たる解析の手段として用い、最新の各種技術と知識を駆使して、複数の研究者による総合的研究を行うものである。本年度は研究代表者を含めて計12名によってこれらの目的を達成することを目標に研究が遂行され、以下に述べる成果が挙げられた。 1.リン脂質平面膜法の一つであるTipーDip法に改良を加え、効率よく膜機能分子を組み込む方法を考案し(高木)、あるいはリン脂質平面膜法を精製のための活性測定の方法として用い、筋小胞体からK^+チャンネル、Cl^ーチャンネルの精製に成功した(葛西)。また、MaxiK^+チャンネルについて、そのイオン透過部位の構造と電位降下部位の推定をリン脂質平面膜法と種々の薬剤を用いることにより解析した(曽我部)。 2.陽イオン性脂質を用いた平面膜の形成(桐野)、両親媒性物質による単分子膜の性質の解析(木下)などからリン脂質平面膜の概念のシミュレ-ションが行われた。 3.特異的に膜融合を促進するペプチドの合成を行った(高橋)。 4.タコ視細胞の光情報伝達に関与すると思われるcGMP結合タンパク質の性質の解析を行い(津田)、cGMPによって活性化されるカチオンチャンネルの存在が示された(平田)。 5.バクテリオロドプシンを平面膜に効率よく再構成する方法が開発された(宗行)。 6.電子顕微鏡を用いてCl^ーチャンネルが12nm×16nmのタマゴ型をしていることがしめされ(藤墳)、無染色、無固定のままで急速凍結を行うクライオ電子顕微鏡法によりタンパク質ー分子の動きを10msオ-ダ-で解析できるようになった(徳永)。 7.リン脂質平面膜の中でのリン脂質分子の動きをコンピュ-タ-シミュレ-ションによって可視化し、リン脂質分子の拡散速度やO_2の透過速度の推定を行った(楠見)。

Research Products
(35 results)

All Other

All Publications (35 results)

[Publications] Kazuki Nunogaki: "How to separate the H^+ and OH^ー permeabilities: Kinetics of intravesicular pH change after medium pH jump." J.Theor.Biol.144. 415-427 (1990)
[Publications] Nobuaki Maeda: "Structural and functional characterization of inositol 1,4,5ーtriphosphate receptor channel from mouse cerebellum." J.Biol.Chem.266. 1109-1116 (1991)
[Publications] Kiyoshi Okihara: "Ca^<2+> and voltageーdependent and calmodulinーantagonistーsensitive anion channel in Chara plasmalemma: A patch clamp study." Protoplasma.
[Publications] 葛西道生: "生体電気信号の発生機構" 学術月報. 43. 124-129 (1990)
[Publications] M.Higuchi: "A simple method for the preparation of an amphiphilic sequential polypeptide." Macromolecules. 23. 361-365 (1990)
[Publications] M.Higuchi: "Channel forming activity of an amphiphilic sequential polypeptide in a cationic bilayer membrane." Bull.Chem.Soc.Jpn.63. 1916-1920 (1990)
[Publications] M.Higuchi: "Interaction between Lーglutamic acid containing polypeptide and Cu^<2+> ion on a bilayer membrane." Senーi Gakkaishi. 46. 384-390 (1990)
[Publications] Y.Nagata: "Copper ion complex formation with a dicarboxylic acidーcontaing polypeptide." Rept.Prog.Polym.Phys.,Japan. 33. 555-558 (1990)
[Publications] M.Higuchi: "Interaction between an anionic amphiphilic sequential polypeptide polypeptide and an anionic bilayer membrane." Polym.J.23. 15-22 (1991)
[Publications] Y.Kirino: "Patchーclamp measurement of ion channels in biomembrane vesicles reconstituted into giant proteoliposomes by freezeーthawing without use of detergent." Methods in Neuroscience.
[Publications] N.Hirashima: "Comparative electrophysiological study of reconstituted giant vesicle preparations of the rabbit skeletal muscle sarcoplasmic reticulam K channel." Biochim.Biophys.Acta.
[Publications] K.Anzai: "Formation of ion channels by synthetic basic peptides in planar lipid bilayer membrane." Biochim.Biophys.Acta.
[Publications] 桐野豊: "骨格筋の細胞内カルシウム動態" 日本臨床(別冊特集:骨格筋疾患). 47. 41-48 (1990)
[Publications] M.Sokabe: "Elecrophysiolgical analysis of structural aspects of voltageーdependent SR K channel." Comp.Biochem.Physiol.
[Publications] M.Sokabe: "Quantitative videomicroscopy of patch calmped membranes: stress,strain,capacitance,and stretch channel activation." Biophys.J.
[Publications] M.Sokabe: "The structure and dynamics of patch clamped membranes: A study by using differential interference light microscopy." J.Cell Biol.111. 599-606 (1990)
[Publications] K.Nomura: "Aminoglycoside blockade of Ca activated K channel from rat brain synaptosomal membranes incorporated into planar bilayers." J.Membr.Biol.115. 241-251 (1990)
[Publications] F.Sachs: "Stretch activated ion channels and membrane mechanics." Neurosci.Res.12. S1-4 (1990)
[Publications] F.Ito: "Effects of ions and drugs on the responses of sensory axon terminals of decapsulated frog muscle spindles." Neurosci.Res.12. S15-26 (1990)
[Publications] C.Fujitsuka: "Intramembrane particles and responses of sensory axon terminals during reinnervation of the frog muscle spindle." J.Neurocytology. 19. 175-185 (1990)
[Publications] A.Yoshimura: "Classification of the intrafusal muscle fibres in the frog muscle spindle: Histochemical and immunohistochemical studies." J.Anatomy(London).
[Publications] 曽我部正博: "筋紡錘感覚神経終末では機械的変形をどのように電気的信号に変換しているか。" 膜(Membrane). 15. 101-109 (1990)
[Publications] S.Takahashi: "Conformation of membrane fusionーactive 20ーresidue peptides with or without lipid bilayers.Implication of alphaーhelix formation for membrane fusion." Biochemistry. 29. 6257-6264 (1990)
[Publications] S.Asakura: "Hydrophobic residues 382ー386 of antithrombin III,AlaーAlaーAlaーSerーThr,serve as the epitope for an antibody which facilitates hydrolysis of the inhibitor by thrombin." J.Biol.Chem.265. 5135-5138 (1990)
[Publications] M.Murata: "Membrane fusion and lysis by the amphiphilic peptides induced by charge neutralization: A model study of virus membrane fusion memchanism." Biophysics of the Cell Surface. 221-236 (1990)
[Publications] M.Tsuda: "Cyclic nucleotides and GTP analogues stimulated lightーinduced phosphorylation of octopus rhodopsin." FEBS Lett.254. 38-40 (1990)
[Publications] M.Tsuda: "Two distinct light regulated Gーproteins in octopus photoreceptors." Biochim.Biophys.Acta. 1052. 204-210 (1990)
[Publications] M.Tokunaga: "Structure and structural change of the myosin head." Advances in Biophysics.
[Publications] M.Okada: "Aromatic rings of tyrosine residues at adenine nucleotide binding sites of the β subunits of F1ーATPase are not necessary for ATPase Activity." Biochem.Biophys.Res.Commun.168. 372-378 (1990)
[Publications] 葛西道生: "イオン透過とイオノフォア(生体膜の分子素子・分子機械)" 学会出版センタ-, 25 (1990)
[Publications] 桐野豊: "“機能再構成(イオンチャンネル)"「レセプタ-」(井村裕夫、岡哲雄、芳賀達也、岸本英爾編)印刷中" 朝倉書店,
[Publications] 桐野豊: "エッセンスESR:基礎と実験" 広川書店,
[Publications] M.Sokabe: "Towards molecular mechanism of the gating in stretch activated channel.in“Comparative Aspect of Mechanoーreception"(ed.F.Ito) in press." Springer Verlag,
[Publications] M.Hasegawa: "Characters and biological behavior of biocompatible dextranーironoxide magnetic fluid.in“Clinical Inplant Materials"" Elsevier,AmsterdamーOxfordーNew YorkーTokyo, 5 (1990)
[Publications] 津田基之: "生命科学の基礎6「生体膜の分子素子機械」" 学会出版, 17 (1990)

1990 Fiscal Year Annual Research Report

生体膜機能分子のリン脂質平面膜法による解析

Principal Investigator

平田 肇 自治医科大学, 医学部, 講師 (40049052)

Research Products

[Publications] Kazuki Nunogaki: "How to separate the H^+ and OH^ー permeabilities: Kinetics of intravesicular pH change after medium pH jump." J.Theor.Biol.144. 415-427 (1990)

[Publications] Nobuaki Maeda: "Structural and functional characterization of inositol 1,4,5ーtriphosphate receptor channel from mouse cerebellum." J.Biol.Chem.266. 1109-1116 (1991)

[Publications] Kiyoshi Okihara: "Ca^<2+> and voltageーdependent and calmodulinーantagonistーsensitive anion channel in Chara plasmalemma: A patch clamp study." Protoplasma.

[Publications] 葛西 道生: "生体電気信号の発生機構" 学術月報. 43. 124-129 (1990)

[Publications] M.Higuchi: "A simple method for the preparation of an amphiphilic sequential polypeptide." Macromolecules. 23. 361-365 (1990)

[Publications] M.Higuchi: "Channel forming activity of an amphiphilic sequential polypeptide in a cationic bilayer membrane." Bull.Chem.Soc.Jpn.63. 1916-1920 (1990)

[Publications] M.Higuchi: "Interaction between Lーglutamic acid containing polypeptide and Cu^<2+> ion on a bilayer membrane." Senーi Gakkaishi. 46. 384-390 (1990)

[Publications] Y.Nagata: "Copper ion complex formation with a dicarboxylic acidーcontaing polypeptide." Rept.Prog.Polym.Phys.,Japan. 33. 555-558 (1990)

[Publications] M.Higuchi: "Interaction between an anionic amphiphilic sequential polypeptide polypeptide and an anionic bilayer membrane." Polym.J.23. 15-22 (1991)

[Publications] Y.Kirino: "Patchーclamp measurement of ion channels in biomembrane vesicles reconstituted into giant proteoliposomes by freezeーthawing without use of detergent." Methods in Neuroscience.

[Publications] N.Hirashima: "Comparative electrophysiological study of reconstituted giant vesicle preparations of the rabbit skeletal muscle sarcoplasmic reticulam K channel." Biochim.Biophys.Acta.

[Publications] K.Anzai: "Formation of ion channels by synthetic basic peptides in planar lipid bilayer membrane." Biochim.Biophys.Acta.

[Publications] 桐野 豊: "骨格筋の細胞内カルシウム動態" 日本臨床(別冊特集:骨格筋疾患). 47. 41-48 (1990)

[Publications] M.Sokabe: "Elecrophysiolgical analysis of structural aspects of voltageーdependent SR K channel." Comp.Biochem.Physiol.

[Publications] M.Sokabe: "Quantitative videomicroscopy of patch calmped membranes: stress,strain,capacitance,and stretch channel activation." Biophys.J.

[Publications] M.Sokabe: "The structure and dynamics of patch clamped membranes: A study by using differential interference light microscopy." J.Cell Biol.111. 599-606 (1990)

[Publications] K.Nomura: "Aminoglycoside blockade of Ca activated K channel from rat brain synaptosomal membranes incorporated into planar bilayers." J.Membr.Biol.115. 241-251 (1990)

[Publications] F.Sachs: "Stretch activated ion channels and membrane mechanics." Neurosci.Res.12. S1-4 (1990)

[Publications] F.Ito: "Effects of ions and drugs on the responses of sensory axon terminals of decapsulated frog muscle spindles." Neurosci.Res.12. S15-26 (1990)

[Publications] C.Fujitsuka: "Intramembrane particles and responses of sensory axon terminals during reinnervation of the frog muscle spindle." J.Neurocytology. 19. 175-185 (1990)

[Publications] A.Yoshimura: "Classification of the intrafusal muscle fibres in the frog muscle spindle: Histochemical and immunohistochemical studies." J.Anatomy(London).

[Publications] 曽我部 正博: "筋紡錘感覚神経終末では機械的変形をどのように電気的信号に変換しているか。" 膜(Membrane). 15. 101-109 (1990)

[Publications] S.Takahashi: "Conformation of membrane fusionーactive 20ーresidue peptides with or without lipid bilayers.Implication of alphaーhelix formation for membrane fusion." Biochemistry. 29. 6257-6264 (1990)

[Publications] S.Asakura: "Hydrophobic residues 382ー386 of antithrombin III,AlaーAlaーAlaーSerーThr,serve as the epitope for an antibody which facilitates hydrolysis of the inhibitor by thrombin." J.Biol.Chem.265. 5135-5138 (1990)

[Publications] M.Murata: "Membrane fusion and lysis by the amphiphilic peptides induced by charge neutralization: A model study of virus membrane fusion memchanism." Biophysics of the Cell Surface. 221-236 (1990)

[Publications] M.Tsuda: "Cyclic nucleotides and GTP analogues stimulated lightーinduced phosphorylation of octopus rhodopsin." FEBS Lett.254. 38-40 (1990)

[Publications] M.Tsuda: "Two distinct light regulated Gーproteins in octopus photoreceptors." Biochim.Biophys.Acta. 1052. 204-210 (1990)

[Publications] M.Tokunaga: "Structure and structural change of the myosin head." Advances in Biophysics.

[Publications] M.Okada: "Aromatic rings of tyrosine residues at adenine nucleotide binding sites of the β subunits of F1ーATPase are not necessary for ATPase Activity." Biochem.Biophys.Res.Commun.168. 372-378 (1990)

[Publications] 葛西 道生: "イオン透過とイオノフォア(生体膜の分子素子・分子機械)" 学会出版センタ-, 25 (1990)

[Publications] 桐野 豊: "“機能再構成(イオンチャンネル)"「レセプタ-」(井村裕夫、岡哲雄、芳賀達也、岸本英爾編)印刷中" 朝倉書店,

[Publications] 桐野 豊: "エッセンスESR:基礎と実験" 広川書店,

[Publications] M.Sokabe: "Towards molecular mechanism of the gating in stretch activated channel.in“Comparative Aspect of Mechanoーreception"(ed.F.Ito) in press." Springer Verlag,

[Publications] M.Hasegawa: "Characters and biological behavior of biocompatible dextranーironoxide magnetic fluid.in“Clinical Inplant Materials"" Elsevier,AmsterdamーOxfordーNew YorkーTokyo, 5 (1990)

[Publications] 津田 基之: "生命科学の基礎6「生体膜の分子素子機械」" 学会出版, 17 (1990)

平田肇自治医科大学, 医学部, 講師 (40049052)

[Publications] 葛西道生: "生体電気信号の発生機構" 学術月報. 43. 124-129 (1990)

[Publications] 桐野豊: "骨格筋の細胞内カルシウム動態" 日本臨床(別冊特集:骨格筋疾患). 47. 41-48 (1990)

[Publications] 曽我部正博: "筋紡錘感覚神経終末では機械的変形をどのように電気的信号に変換しているか。" 膜(Membrane). 15. 101-109 (1990)

[Publications] 葛西道生: "イオン透過とイオノフォア(生体膜の分子素子・分子機械)" 学会出版センタ-, 25 (1990)

[Publications] 桐野豊: "“機能再構成(イオンチャンネル)"「レセプタ-」(井村裕夫、岡哲雄、芳賀達也、岸本英爾編)印刷中" 朝倉書店,

[Publications] 桐野豊: "エッセンスESR:基礎と実験" 広川書店,

[Publications] 津田基之: "生命科学の基礎6「生体膜の分子素子機械」" 学会出版, 17 (1990)