1989 年度実績報告書

異環境生物のエネルギー

研究課題

研究課題/領域番号	01617003
研究機関	名古屋大学
研究代表者	向畑恭男名古屋大学, 理学部, 教授 (10028110)
研究分担者	土屋友房岡山大学, 薬学部, 教授 (80012673) 今栄康雄名古屋大学, 理学部, 助教授 (70022712) 前島正義北海道大学, 低温科学研究所, 助手 (80181577) 山中健生東京工業大学, 理学部, 教授 (40028113) 吉田賢右東京工業大学, 理学部, 教授 (90049073)
キーワード	ハロバクテリア / スルホロブス / 鉄酸化細菌 / 液胞 / アーキロドプシン / H^+ ATPアーゼ / H^+ PPiアーゼ / チトクロム酸化酵素
研究概要	この研究では、異常な環境に生息する微生物が工夫/改変して持つ独特のエネルギー転換系の構造と作動機作を解明することと、これを通して常環境の生物のエネルギー転換系についての理解を深めることを目的とした。本年度研究結果抜粋:高度好塩性細菌ハロバクテリアの新種からバクテリオロドプシン(bR)とも、前年度に発見のアーキロドプシン(aR)とも異なる光駆動H^+ポンプを同定し、アーキロドプシンー2と命名した。この新レチナールタンパク質の遺伝子をクローン化して、DNA塩基/アミノ酸配列を決めたところ、bRとの相同性は60%弱、aRとは90%弱であり、3者で特にH^+の通路を形成するC、F、Gの3本のヘリックスのアミノ酸の保存度が高かった。これらの結果からポンプタンパク質の必須立体構造についての多くの重要な示唆を得た。さらにATPアーゼ遺伝子をクローン化して、主2サブユニットのDNA塩基/アミノ酸配列を決定した。この配列は前年度に決定した好酸好熱性古細菌スルホロブスとメタン産生古細菌のATPアーゼ、および液胞のV型ATPアーゼのいずれのアミノ酸配列とも相同性が高く、ミトコンドリアのF型ATPアーゼのそれとは低くて、初年度に免疫交差反応で得た新知見が確証された。スルホロブスATPアーゼでは同一オペロン内の全遺伝子のDNA塩基/アミノ酸配列を決定し、ホロ酵素が、少なくとも膜内に1種、膜上に5種のサブユニットを持つと推定した。好酸性鉄酸化細菌から、前年度に同定したpH3を至適とするCyt552と対となる酸化酵素を精製して酵素化学的性質を精査し、この酵素が従来の予測とは逆のペリプラズマ側で機能していると推定した。植物液胞膜からH^+輸送性で73kDaのピロフォスファターゼを初めて単離精製した。この酵素は分子量、抗原性等で従来知られたものと異なることを見出した。これらの結果は極めて斬新なものである。

研究成果
(22件)

すべてその他

すべて文献書誌 (22件)

[文献書誌] Yasuo Sugiyama: "Isolation of a gene that encodes a new retinal protein, Archaerhodopsin,from Halobacterium sp.aus-1" Journal of Bilogical Chemistry. 264. 20859-20862 (1989)
[文献書誌] Yasuo Mukohata: "Close evolutionary relatedness of archaebacteria with eukaryotes" Proceedings of Japan Academy. (1990)
[文献書誌] Kunio Ihara: "The amino acid sequences of the and subunits of ATP synthase of Halobacterium halobium" Journal of Biological Chemistry. (1990)
[文献書誌] Koichi Uegaki: "The amino acid sequence of Archaerhodopsin-2,Australian bacteriorhodopsin-like protein" Archives of Biochemistry and Biophysics. (1990)
[文献書誌] Yasuo Mukohata: "Australian halobacteria and their ion-pumping rhodopsins" Photochemistry and Photobiology. (1990)
[文献書誌] Yasuo Imae: "Na^+-driven bacterial flagellar motors" Journal of Bioenergetics and Biomembranes. 21. 705-716 (1989)
[文献書誌] Takashi Atsumi: "Specific inhibition of the Na^+-driven flagellar motors of alkalophilic Bacillus strains by the amiloride analog phenamil" Journal of Bacteriology. 172. (1990)
[文献書誌] Naoki Kamo: "Linear equation for calibration of the mitochondrial membrane potential" Bioelectrochemistry and Bioenergetics. 21. 33-45 (1989)
[文献書誌] Shigeru Kurosawa: "Plasma-polymerized membrane electrode selective for lopophilic cations" Journal of Membrane Science. 46. 157-166 (1989)
[文献書誌] Takao Kodama: "A stopped-flow calorimeter with a thermocouple array sensor for studies of energetic aspects of enzymatic reactions" Thermochimica Acta. (1990)
[文献書誌] Yuki Sakai: "A respiratory-driven and an artificially-driven ATP synthesis in mutants of Vibrio parahaemolyticus" Biochimica et Biophysica Acta. 973. 450-456 (1989)
[文献書誌] Chie Moritani: "Characteristics of the H^+translocating ATPase of Vibrio parahaemolyticus" Chemical and Pharmaceutical Bulletin. 38. 164-167 (1990)
[文献書誌] Kazuhiro Kogure: "Respiration-dependent primary Na^+ pump in halophilic marine bacterium,Alcaligenes strain 201" FEBS Letters. 256. 147-149 (1989)
[文献書誌] Naoto Murakami: "Mechanism of function of dicyclohexylcarbodiimide-sensitive Na^+/H^+ antiporter in H__ー.halobium:pH effect" Archives of Biochemistry and Biophysics. 271. 515-523 (1989)
[文献書誌] Masayoshi Maeshima: "Purification and properties of vacuolar membrane proton translocating inorganic pyrophosphatase from mung bean" Journal of Biological Chemistry. 264. 20068-20073 (1989)
[文献書誌] Chie Matsuura-Endo: "Subunit composition of vacuolar membrane H^+-ATPase from mung bean" European Journal of Biochemistry. (1990)
[文献書誌] Masahiro Kai: "Cytochrome oxidase of an acidophilic iron-oxidizing bacterium,Thiobacillus ferrooxidans,functions at pH 3.5" Biochemical and Biophysical Research Communication. 160. 839-843 (1989)
[文献書誌] Akihiko Sato: "Thiobacillus ferroxidans cytochrome c__ー-552:Purification and some of its molecular features" Biochimica et Biophysica Acta. 976. 129-134 (1989)
[文献書誌] Kiminori Denda: "A gene encoding the proteolipid subunit of Sulfolobus acidocaldarius ATPase complex" Journal of Biological Chemistry. 264. 7119-7121 (1989)
[文献書誌] J.P.Gogarten: "The evolution of the vacuolar H^+-ATPase:Implication for origin of eukaryotes" Proceedings of National Academy of Science,USA. 86. 6661-6665 (1989)
[文献書誌] Ken Yokoyama: "The reconstituted 33 complex of the thermostable F_1-ATPase" Journal of Biological Chemistry. 264. 21837-21841 (1989)
[文献書誌] M.H.Qureshi: "A novel aco-type cytochrome-c__ー oxidase from a facultative alkalophilic Bacillus:Purification,and some molecular and enzymatic features" Journal of Biochemistry. 107. (1990)

1989 年度 実績報告書

異環境生物のエネルギー

研究代表者

向畑 恭男 名古屋大学, 理学部, 教授 (10028110)

研究成果

[文献書誌] Yasuo Sugiyama: "Isolation of a gene that encodes a new retinal protein, Archaerhodopsin,from Halobacterium sp.aus-1" Journal of Bilogical Chemistry. 264. 20859-20862 (1989)

[文献書誌] Yasuo Mukohata: "Close evolutionary relatedness of archaebacteria with eukaryotes" Proceedings of Japan Academy. (1990)

[文献書誌] Kunio Ihara: "The amino acid sequences of the and subunits of ATP synthase of Halobacterium halobium" Journal of Biological Chemistry. (1990)

[文献書誌] Koichi Uegaki: "The amino acid sequence of Archaerhodopsin-2,Australian bacteriorhodopsin-like protein" Archives of Biochemistry and Biophysics. (1990)

[文献書誌] Yasuo Mukohata: "Australian halobacteria and their ion-pumping rhodopsins" Photochemistry and Photobiology. (1990)

[文献書誌] Yasuo Imae: "Na^+-driven bacterial flagellar motors" Journal of Bioenergetics and Biomembranes. 21. 705-716 (1989)

[文献書誌] Takashi Atsumi: "Specific inhibition of the Na^+-driven flagellar motors of alkalophilic Bacillus strains by the amiloride analog phenamil" Journal of Bacteriology. 172. (1990)

[文献書誌] Naoki Kamo: "Linear equation for calibration of the mitochondrial membrane potential" Bioelectrochemistry and Bioenergetics. 21. 33-45 (1989)

[文献書誌] Shigeru Kurosawa: "Plasma-polymerized membrane electrode selective for lopophilic cations" Journal of Membrane Science. 46. 157-166 (1989)

[文献書誌] Takao Kodama: "A stopped-flow calorimeter with a thermocouple array sensor for studies of energetic aspects of enzymatic reactions" Thermochimica Acta. (1990)

[文献書誌] Yuki Sakai: "A respiratory-driven and an artificially-driven ATP synthesis in mutants of Vibrio parahaemolyticus" Biochimica et Biophysica Acta. 973. 450-456 (1989)

[文献書誌] Chie Moritani: "Characteristics of the H^+translocating ATPase of Vibrio parahaemolyticus" Chemical and Pharmaceutical Bulletin. 38. 164-167 (1990)

[文献書誌] Kazuhiro Kogure: "Respiration-dependent primary Na^+ pump in halophilic marine bacterium,Alcaligenes strain 201" FEBS Letters. 256. 147-149 (1989)

[文献書誌] Naoto Murakami: "Mechanism of function of dicyclohexylcarbodiimide-sensitive Na^+/H^+ antiporter in H__ー.halobium:pH effect" Archives of Biochemistry and Biophysics. 271. 515-523 (1989)

[文献書誌] Masayoshi Maeshima: "Purification and properties of vacuolar membrane proton translocating inorganic pyrophosphatase from mung bean" Journal of Biological Chemistry. 264. 20068-20073 (1989)

[文献書誌] Chie Matsuura-Endo: "Subunit composition of vacuolar membrane H^+-ATPase from mung bean" European Journal of Biochemistry. (1990)

[文献書誌] Masahiro Kai: "Cytochrome oxidase of an acidophilic iron-oxidizing bacterium,Thiobacillus ferrooxidans,functions at pH 3.5" Biochemical and Biophysical Research Communication. 160. 839-843 (1989)

[文献書誌] Akihiko Sato: "Thiobacillus ferroxidans cytochrome c__ー-552:Purification and some of its molecular features" Biochimica et Biophysica Acta. 976. 129-134 (1989)

[文献書誌] Kiminori Denda: "A gene encoding the proteolipid subunit of Sulfolobus acidocaldarius ATPase complex" Journal of Biological Chemistry. 264. 7119-7121 (1989)

[文献書誌] J.P.Gogarten: "The evolution of the vacuolar H^+-ATPase:Implication for origin of eukaryotes" Proceedings of National Academy of Science,USA. 86. 6661-6665 (1989)

[文献書誌] Ken Yokoyama: "The reconstituted 33 complex of the thermostable F_1-ATPase" Journal of Biological Chemistry. 264. 21837-21841 (1989)

[文献書誌] M.H.Qureshi: "A novel aco-type cytochrome-c__ー oxidase from a facultative alkalophilic Bacillus:Purification,and some molecular and enzymatic features" Journal of Biochemistry. 107. (1990)

1989 年度実績報告書

向畑恭男名古屋大学, 理学部, 教授 (10028110)