1991 Fiscal Year Annual Research Report

膜タンパク質ス-パ-コンプレックスの構築原理と結晶化条件の解明

Research Project

Project/Area Number	03304056
Research Institution	Himeji Institute of Technology
Principal Investigator	吉川信也姫路工業大学, 理学部, 教授 (40068119)
Co-Investigator(Kenkyū-buntansha)	豊島近東京工業大学, 生命理工学部, 助教授 (70172210) 月原冨武徳島大学, 工学部, 教授 (00032277) 白木原康雄兵庫教育大学, 自然系, 助教授 (20150287) 神山勉理化学研究所, 生物物理, 研究員安宅光雄繊維高分子材料研究所, 生体工学, 主任研究員
Keywords	膜タンパク質 / 結晶化 / 結晶構造解析 / チトクロム酸化酵素 / チトクロムbc_1複合体
Research Abstract	結晶化条件の検討:チトクロム酸化酵素の結晶化条件に対する温度の効果を検討したところ20℃でプラスチック容器でしか析出しない結晶が得られた。このことは容器の材質も結晶化条件の重要な要因であることを示している(吉川)。またウシ心筋から精製したComplexIIIの結晶化にも成功し、6.8Å分解能のX線解析を得た(松原)。F_1ATPaseおよびそのα_3β_3複合体の、それぞれ8Åおよび4Å分解能の反射を示すような結晶が得られるようになった(白木原)。バクテリオロドプシンも分解能は不十分ではあるが、これまでに報告されていない六方晶系の結晶が得られた。またベン毛モ-タMリングの構成タンパク質をコ-ドする遺伝子を発現させ、精製することに成功した。その他、光合成関連膜タンパク質複合体や結晶化条件の検討についても着実な進歩があった。構築原理と結晶成長機構:アミノ酸配列から立体構造の情報を抽出するための計算方法の開発を行っているが、今年度はプロ-ブヘリック入法により膜貫通型ヘリックスの性質をエネルギ-計算により詳細に調べた(美宅)。またチトクロム酸化酵素の結晶化の相図を膜タンパク質としてははじめて決定し、六方晶系の結晶は塩溶域で析出することを示した(安宅)。構造解析:電子線回折による電子密度図を初期値とした分子置換法を行うとき、できるだけ初期値の影響をうけないようにするために計算法を改良し、初期値として重心の位置と楕円近似だけで計算が可能となった(月原)。分子置換法による結果の評価に必要な電子線回折による電子密度の精密化のため、二次元結晶を作成し、20Å分解能程度の回折像は再現性よく得られるようになった(豊島)。研究会:これらの成果は平成4年1月に行われた研究会で発表され、有意義な討論が行われた。

Research Products
(19 results)

All Other

All Publications (19 results)

[Publications] Kubori,T.: "Morphological pathway of Flagellar Assembly in Salmonella typhimurium" J.Mol.Biol.
[Publications] Ueno,T.: "The M ring,S ring and proximal rod of the flagellar basal body of Salmonella typhimurium are composed of subunits of a single protein,FliF." J.Mol.Biol.
[Publications] Ataka,M.: "Phase Diagrams of a Crystalline Membrane Protein,Bovine Heart Cytochrome c Ocidase,in the SaltingーIn Region." J.Cryst.Growth.
[Publications] Elgersma,A.V.: "Kinetic Studies on the Growth of Three Crystal Forms of Lysozyme Based on the Measurement of Protein and Cl^ーConcentraition Changes." J.Cryst.Growth.
[Publications] Nagamune,T.: "Crystallization of a Photosensitive Nitrile Hydratase from Rhodococcus sp.Nー771." J.Mol.Biol.220. 221-222 (1991)
[Publications] Kamiya,N.: "Heme Environmental Structure of a Novel Artificial Myoglobin with Closed Heme Pocket: SiteーSpecific Chemical Modification Producing Distal NーTetrazolylhistidine E7 by Cyanogen Bromide and Azide Ion." J.Am.Chem.Soc.113. 1826-1829 (1991)
[Publications] Sugawara,Y.: "HumidityーControlled Reversible Structure Transition of Disodium Adenosine 5'ーTriphosphate between Dihydrate and Trihydrate in a Single Crystal State." J.Am.Chem.Soc.113. 5440-5445 (1991)
[Publications] Kobayashi,T.: "Femtosecond Spectroscopy of Acidified & Neutral Bacteriorhodopsin." Soc.PhotoーOptic.Instr.Engineer.403. 407-416 (1991)
[Publications] Shirakihara,Y.: "Purification by dyeーligand chromatography and a crystallization study of F_1ーATPase and its major subunits,β and α,from a thermophilic bacterium,PS3" J.Biochem.109. 466-471 (1991)
[Publications] Toyoshima,C.: "Contrast transfer in frozenーhydrated electron microscopy II.Amplitude co ntrast at very low frequencies." Ultramicroscopy.
[Publications] Taki,M.: "Synthesis of Diazipine and[ ^3H]Diazipine:Novel Dihydropyridine for Photoaffinity Probes." Chem.Pharm.Bull.33. 1860-1862 (1991)
[Publications] Taki,M.: "Diazipine,A Novel Photoaffinity Probe for Dihydropyridine Receptors of Calcium Channels." FEBS Lett.283. 259-262 (1991)
[Publications] Nakayama,H.: "Identification of the 1,4ーDihydropyridine Binding Regions within the alpha l Subunit of Skeletal Muscle Ca^<2+> channels by Photoaffinity Labeling with Diazipine." Proc.Natl.Acad.Sci.USA.88. 9203-9207 (1991)
[Publications] Nakayama,H.(H.Glossmann,J.striessnig,eds): "Synthesis and Properties of Diazipine,A Novel Photoaffinity Probe of Calcium ChannelーLinked Dihydropyridine Receptors." Methods in Pharmacology. 7.
[Publications] Fujii,I.: "Evaluation of Xーray Diffraction Data from Protein Crystals with Use of an Imaging Plate." Acta Crystallogr.B47. 137-144 (1991)
[Publications] Kita,A.: "Crystallization and Preliminary Xーray Diffraction Studies of a Flavoprote in,FP_<390> from a Luminescent Bacterium,Photobacteri phosphoreum." J.Biochem.110. 748-750 (1991)
[Publications] Yoshikawa,S.: "Crystallization of beef heart cytochrome c oxidase." Jounal of Crystal Growth.110. 247-251 (1991)
[Publications] Kubota,T.: "Crystallization and Preliminary Xーray Crystallographic Studies of Bovine Heart Mitochondrial Cytochrome bc_1 Complex." J.Mol.Biol.221. 379-382 (1991)
[Publications] Yoshikawa,S.: "Strategies for crystallization of large membrane protein complexes." Journal of Crystal Growth.

1991 Fiscal Year Annual Research Report

膜タンパク質ス-パ-コンプレックスの構築原理と結晶化条件の解明

Principal Investigator

吉川 信也 姫路工業大学, 理学部, 教授 (40068119)

Research Products

[Publications] Kubori,T.: "Morphological pathway of Flagellar Assembly in Salmonella typhimurium" J.Mol.Biol.

[Publications] Ueno,T.: "The M ring,S ring and proximal rod of the flagellar basal body of Salmonella typhimurium are composed of subunits of a single protein,FliF." J.Mol.Biol.

[Publications] Ataka,M.: "Phase Diagrams of a Crystalline Membrane Protein,Bovine Heart Cytochrome c Ocidase,in the SaltingーIn Region." J.Cryst.Growth.

[Publications] Elgersma,A.V.: "Kinetic Studies on the Growth of Three Crystal Forms of Lysozyme Based on the Measurement of Protein and Cl^ーConcentraition Changes." J.Cryst.Growth.

[Publications] Nagamune,T.: "Crystallization of a Photosensitive Nitrile Hydratase from Rhodococcus sp.Nー771." J.Mol.Biol.220. 221-222 (1991)

[Publications] Kamiya,N.: "Heme Environmental Structure of a Novel Artificial Myoglobin with Closed Heme Pocket: SiteーSpecific Chemical Modification Producing Distal NーTetrazolylhistidine E7 by Cyanogen Bromide and Azide Ion." J.Am.Chem.Soc.113. 1826-1829 (1991)

[Publications] Sugawara,Y.: "HumidityーControlled Reversible Structure Transition of Disodium Adenosine 5'ーTriphosphate between Dihydrate and Trihydrate in a Single Crystal State." J.Am.Chem.Soc.113. 5440-5445 (1991)

[Publications] Kobayashi,T.: "Femtosecond Spectroscopy of Acidified & Neutral Bacteriorhodopsin." Soc.PhotoーOptic.Instr.Engineer.403. 407-416 (1991)

[Publications] Shirakihara,Y.: "Purification by dyeーligand chromatography and a crystallization study of F_1ーATPase and its major subunits,β and α,from a thermophilic bacterium,PS3" J.Biochem.109. 466-471 (1991)

[Publications] Toyoshima,C.: "Contrast transfer in frozenーhydrated electron microscopy II.Amplitude co ntrast at very low frequencies." Ultramicroscopy.

[Publications] Taki,M.: "Synthesis of Diazipine and[ ^3H]Diazipine:Novel Dihydropyridine for Photoaffinity Probes." Chem.Pharm.Bull.33. 1860-1862 (1991)

[Publications] Taki,M.: "Diazipine,A Novel Photoaffinity Probe for Dihydropyridine Receptors of Calcium Channels." FEBS Lett.283. 259-262 (1991)

[Publications] Nakayama,H.: "Identification of the 1,4ーDihydropyridine Binding Regions within the alpha l Subunit of Skeletal Muscle Ca^<2+> channels by Photoaffinity Labeling with Diazipine." Proc.Natl.Acad.Sci.USA.88. 9203-9207 (1991)

[Publications] Nakayama,H.(H.Glossmann,J.striessnig,eds): "Synthesis and Properties of Diazipine,A Novel Photoaffinity Probe of Calcium ChannelーLinked Dihydropyridine Receptors." Methods in Pharmacology. 7.

[Publications] Fujii,I.: "Evaluation of Xーray Diffraction Data from Protein Crystals with Use of an Imaging Plate." Acta Crystallogr.B47. 137-144 (1991)

[Publications] Kita,A.: "Crystallization and Preliminary Xーray Diffraction Studies of a Flavoprote in,FP_<390> from a Luminescent Bacterium,Photobacteri phosphoreum." J.Biochem.110. 748-750 (1991)

[Publications] Yoshikawa,S.: "Crystallization of beef heart cytochrome c oxidase." Jounal of Crystal Growth.110. 247-251 (1991)

[Publications] Kubota,T.: "Crystallization and Preliminary Xーray Crystallographic Studies of Bovine Heart Mitochondrial Cytochrome bc_1 Complex." J.Mol.Biol.221. 379-382 (1991)

[Publications] Yoshikawa,S.: "Strategies for crystallization of large membrane protein complexes." Journal of Crystal Growth.

吉川信也姫路工業大学, 理学部, 教授 (40068119)