1994 年度実績報告書

タンパク質折り畳み中間体の構造と構造形成機構

研究課題

研究課題/領域番号	06304051
研究機関	大阪大学
研究代表者	片岡幹雄大阪大学, 理学部, 助教授 (30150254)
研究分担者	木原裕関西医科大学, 医学部, 教授 (20049076) 月向邦彦名古屋大学, 農学部, 助教授 (10023467) 桑島邦博東京大学大学院, 理学研究科, 助教授 (70091444) 赤坂一之神戸大学, 理学部, 教授 (50025368) 後藤祐児大阪大学, 理学部, 助教授 (40153770)
キーワード	蛋白質折り畳み / モルテングロビュール / 変性 / X線溶液散乱 / ストップトフローCD / 静電的相互作用 / 疎水的相互作用 / 蛋白質非天然構造
研究概要	アポミオグロビンやチトクロムcのモルテングロビュール状態の溶液構造がX線溶液散乱法により詳細に比較され、モルテングロビュール構造は、サイズ、形状、内部構造などにより二通りに分類されることがわかった。一方は、天然構造に近く、他は変性構造に近い。後者は、疎水的な核と揺動する部分からなっていると考えられる。天然構造がβ構造のみからなるβラクトグロブリンは、有機溶媒中でαヘリックス構造をとる。また、βラクトグロブリンは二次構造予測によれば、α蛋白と予測される。有機溶媒中のヘリックス含量は、予測されるヘリックス含量とよい相関を示した。こうしたことから、構造形成の途上で現れる中間状態には天然の構造とは全く異なる二次構造ができているという非階層構造形成モデルもありうることが示唆された。トリプトファン合成酵素αサブユニットの変性剤変性は3状態変性、また熱変性は2状態変性であることが示された。変性剤変性の中間状態は、これまで1ドメインがインタクトで他のドメインが変性しているとされていたが、モルテングロビュール様であることが判明した。スタフィロコッカル・ヌクレアーゼの様々な変異体を用いた巻戻り及び変性の速度論的解析がストップトフローCD法により詳細に調べられた。その結果、巻き戻りの遷移状態では天然状態類似の構造が形成されていること、天然状態の安定性や巻戻り速度には変性状態での水和自由エネルギーの寄与が重要であることが示された。蛋白質の折り畳み中間体に関する熱力学的研究が行われ、クーロン相互作用の寄与や比熱の変化が予測された。折り畳み過程の統計力学的研究により、折り畳みの物理的描像が示された。

研究成果
(28件)

すべてその他

すべて文献書誌 (28件)

[文献書誌] Ichiro Nishii: "Cold-denaturation of the molten globule state of apomyoglobin and a profile of protein folding" Biochemistry. 33. 4903-4909 (1994)
[文献書誌] Mikio Kataoka: "Energy coupling in an ion pump:The reprotonation switch of bacteriorhodopsin" Journal of Molecular Biology. 243. 621-638 (1994)
[文献書誌] Mikio Kataoka: "Structural characterization of molten globule and native stares of apomyoglobin by solution X-ray scattering" Journal of Molecular Biology. (印刷中). (1995)
[文献書誌] Yoshihisa Hagihara: "Comparison of the conformational stability of the molten globule and native states of horse cytochrome c:Effects of acetylation,heat,urea and guanidine-hydrochloride" Journal of Molecular Biology. 237. 336-348 (1994)
[文献書誌] Daizo Hamada: "Salt-induced formation of the molten globule state of cytochrome c studied by isothermal titration calorimetry" Proceedings of National Academy of Science U.S.A.91. 10325-10329 (1994)
[文献書誌] Yohihisa Hagihara: "Thermal unfolding of tetrameric melittin:Comparison with the molten globule state of cytochrome c" Protein Science. 3. 1418-1429 (1994)
[文献書誌] Masaru Hoshino: "Perchlorate-induced formation ofthe α-helical structure of mastoparan" Journal of Biochemistry. 116. 910-915 (1994)
[文献書誌] Kentaro Shiraki: "Trifluoroethanol-induced stabilizaytion of the α-helical structure of β-lactoglobulin:Implication for non-hierarchical protein folding" Journal of Molecular Biology. 245. 180-194 (1995)
[文献書誌] Kouhei Tsumoto: "Contribution to antibody-antigen interaction of structurally purterbed anatigenetic residues upon antibody binding" Journal of Biological Chemistry. 269. 28777-28782 (1994)
[文献書誌] Kouhei Tsumoto: "Effect of the order of antibody variable regions on the expression of the single-chain hyhe110 FV fragment in E,coli and the thermodynamic analysis of its antigen binding properities" Biochemical Biophysical Research Communication. 201. 546-551 (1994)
[文献書誌] M.Odaka: "Tyr-341 of the beta subunit is a major Km-determining residue of TF1-ATPase:Parallel effect of its mutations on Kd(ATP)of the β subunit and on Km(ATP)of the α3β3γ complex" Journal of Biochemistry. 115. 789-796 (1994)
[文献書誌] Kyoko Ogasahara: "Unfolding-refolding kinetics of the tryptophan synthase alpha subunit by the CD and fluorescence measurements" Journal of Molecular Biology. 236. 1227-1240 (1994)
[文献書誌] Kazuyuki Akasaka: "Local order,conformation,and interaction in nematic 4-n-pentyloxy-4'-cyanobiphenyl and its one-to-one mixture with 1-(4'-cyanophenyl)-4-propylcyclohexane" Journal of Physical Chemistry. (印刷中). (1995)
[文献書誌] Kazuyuki Akasaka: "Three-dimensional structure of gurumarin,a sweet taste-suppressing polypeptide" Jounal of Biomolecular NMR. (印刷中). (1995)
[文献書誌] Hideo Arakawa: "Conformational changes of α-2 macroglobulin determined by time resolved X-ray solution scattering" FEBS Letters. 337. 171-174 (1994)
[文献書誌] Kaori Chiba: "Kinetic study on myoglobin refolding monitored by five optical stopped-flow methods" Proteins. 19. 110-119 (1994)
[文献書誌] Hiroshi Kihara: "Stoped-flow apparatus for X-ray scattering and XAFS" Journal of Synchrotron Radiation. 1. 74-77 (1994)
[文献書誌] Hirotsugu Tsuruta: "Kinetics of the quartemary structure change of aspartate transcarbamylase triggered by succinate a competitive inhibitor" Biochemistry. 33. 10007-10012 (1994)
[文献書誌] Yoshihiko Igarashi: "Solution X-ray scattering study on the chaperonin GroEL for Escherichia coli" Biophysical Chemistry. 53. 259-266 (1995)
[文献書誌] Akira Okazaki: "The chaperonin GroEL does not recognize apo-α-lactalbumin in the molten globule state" Nature:Structural Biology. 1. 439-446 (1994)
[文献書誌] 桑島邦博: "折れ畳み機構" 蛋白質核酸酵素. 39. 1011-1016 (1994)
[文献書誌] Kunihiko Gekko: "Point mutations at glycine-121 of Escherichia coli dehydrofolate reductase:Important roles of a flexible loop in the stability and function" Journal of Biochemistry. 116. 34-41 (1994)
[文献書誌] Yoichi Tamura: "Compactness of thermally and chemically denatured ribonclease A as revealed by volume and compressibility" Biochemistry. 34. 1878-1884 (1994)
[文献書誌] 月向邦彦: "高圧下における食品高分子のゾル-ゲル転移" New Food Industry. 36. 68-79 (1994)
[文献書誌] Kunitsugu Soda: "Transfer-thermodynamic quantities and the hydrophobic effect" Journal of Physical Society of Japan. 63. 814- (1994)
[文献書誌] 曽田邦嗣: "蛋白質の構造安定性の熱力学と疎水効果" ぶんせき. 4(印刷中). (1995)
[文献書誌] Shuichi Hirono: "Non-congeneric structure-pharmacokinetic property correlation studies using fuzzy adaptive least-squares″:Volume of distribution" Biol.Pharm.Bulletin. 17. 686-690 (1994)
[文献書誌] H.Tsujishita: "Potential-scaled/hot-solute molecular dynamics for modeling of the three-demensional structure of polypeptides in solution" Biophysical Journal. 66. 1815-1822 (1994)

1994 年度 実績報告書

タンパク質折り畳み中間体の構造と構造形成機構

研究代表者

片岡 幹雄 大阪大学, 理学部, 助教授 (30150254)

研究成果

[文献書誌] Ichiro Nishii: "Cold-denaturation of the molten globule state of apomyoglobin and a profile of protein folding" Biochemistry. 33. 4903-4909 (1994)

[文献書誌] Mikio Kataoka: "Energy coupling in an ion pump:The reprotonation switch of bacteriorhodopsin" Journal of Molecular Biology. 243. 621-638 (1994)

[文献書誌] Mikio Kataoka: "Structural characterization of molten globule and native stares of apomyoglobin by solution X-ray scattering" Journal of Molecular Biology. (印刷中). (1995)

[文献書誌] Yoshihisa Hagihara: "Comparison of the conformational stability of the molten globule and native states of horse cytochrome c:Effects of acetylation,heat,urea and guanidine-hydrochloride" Journal of Molecular Biology. 237. 336-348 (1994)

[文献書誌] Daizo Hamada: "Salt-induced formation of the molten globule state of cytochrome c studied by isothermal titration calorimetry" Proceedings of National Academy of Science U.S.A.91. 10325-10329 (1994)

[文献書誌] Yohihisa Hagihara: "Thermal unfolding of tetrameric melittin:Comparison with the molten globule state of cytochrome c" Protein Science. 3. 1418-1429 (1994)

[文献書誌] Masaru Hoshino: "Perchlorate-induced formation ofthe α-helical structure of mastoparan" Journal of Biochemistry. 116. 910-915 (1994)

[文献書誌] Kentaro Shiraki: "Trifluoroethanol-induced stabilizaytion of the α-helical structure of β-lactoglobulin:Implication for non-hierarchical protein folding" Journal of Molecular Biology. 245. 180-194 (1995)

[文献書誌] Kouhei Tsumoto: "Contribution to antibody-antigen interaction of structurally purterbed anatigenetic residues upon antibody binding" Journal of Biological Chemistry. 269. 28777-28782 (1994)

[文献書誌] Kouhei Tsumoto: "Effect of the order of antibody variable regions on the expression of the single-chain hyhe110 FV fragment in E,coli and the thermodynamic analysis of its antigen binding properities" Biochemical Biophysical Research Communication. 201. 546-551 (1994)

[文献書誌] M.Odaka: "Tyr-341 of the beta subunit is a major Km-determining residue of TF1-ATPase:Parallel effect of its mutations on Kd(ATP)of the β subunit and on Km(ATP)of the α3β3γ complex" Journal of Biochemistry. 115. 789-796 (1994)

[文献書誌] Kyoko Ogasahara: "Unfolding-refolding kinetics of the tryptophan synthase alpha subunit by the CD and fluorescence measurements" Journal of Molecular Biology. 236. 1227-1240 (1994)

[文献書誌] Kazuyuki Akasaka: "Local order,conformation,and interaction in nematic 4-n-pentyloxy-4'-cyanobiphenyl and its one-to-one mixture with 1-(4'-cyanophenyl)-4-propylcyclohexane" Journal of Physical Chemistry. (印刷中). (1995)

[文献書誌] Kazuyuki Akasaka: "Three-dimensional structure of gurumarin,a sweet taste-suppressing polypeptide" Jounal of Biomolecular NMR. (印刷中). (1995)

[文献書誌] Hideo Arakawa: "Conformational changes of α-2 macroglobulin determined by time resolved X-ray solution scattering" FEBS Letters. 337. 171-174 (1994)

[文献書誌] Kaori Chiba: "Kinetic study on myoglobin refolding monitored by five optical stopped-flow methods" Proteins. 19. 110-119 (1994)

[文献書誌] Hiroshi Kihara: "Stoped-flow apparatus for X-ray scattering and XAFS" Journal of Synchrotron Radiation. 1. 74-77 (1994)

[文献書誌] Hirotsugu Tsuruta: "Kinetics of the quartemary structure change of aspartate transcarbamylase triggered by succinate a competitive inhibitor" Biochemistry. 33. 10007-10012 (1994)

[文献書誌] Yoshihiko Igarashi: "Solution X-ray scattering study on the chaperonin GroEL for Escherichia coli" Biophysical Chemistry. 53. 259-266 (1995)

[文献書誌] Akira Okazaki: "The chaperonin GroEL does not recognize apo-α-lactalbumin in the molten globule state" Nature:Structural Biology. 1. 439-446 (1994)

[文献書誌] 桑島邦博: "折れ畳み機構" 蛋白質核酸酵素. 39. 1011-1016 (1994)

[文献書誌] Kunihiko Gekko: "Point mutations at glycine-121 of Escherichia coli dehydrofolate reductase:Important roles of a flexible loop in the stability and function" Journal of Biochemistry. 116. 34-41 (1994)

[文献書誌] Yoichi Tamura: "Compactness of thermally and chemically denatured ribonclease A as revealed by volume and compressibility" Biochemistry. 34. 1878-1884 (1994)

[文献書誌] 月向邦彦: "高圧下における食品高分子のゾル-ゲル転移" New Food Industry. 36. 68-79 (1994)

[文献書誌] Kunitsugu Soda: "Transfer-thermodynamic quantities and the hydrophobic effect" Journal of Physical Society of Japan. 63. 814- (1994)

[文献書誌] 曽田邦嗣: "蛋白質の構造安定性の熱力学と疎水効果" ぶんせき. 4(印刷中). (1995)

[文献書誌] Shuichi Hirono: "Non-congeneric structure-pharmacokinetic property correlation studies using fuzzy adaptive least-squares″:Volume of distribution" Biol.Pharm.Bulletin. 17. 686-690 (1994)

[文献書誌] H.Tsujishita: "Potential-scaled/hot-solute molecular dynamics for modeling of the three-demensional structure of polypeptides in solution" Biophysical Journal. 66. 1815-1822 (1994)

1994 年度実績報告書

片岡幹雄大阪大学, 理学部, 助教授 (30150254)