1990 Fiscal Year Annual Research Report

H^+ーATPaseおよびH^+/K^+ ATPaseによるイオン輸送機構の解析

Research Project

Project/Area Number	01480528
Research Institution	Osaka University
Principal Investigator	二井將光大阪大学, 産業科学研究所, 教授 (50012646)
Co-Investigator(Kenkyū-buntansha)	森山芳則大阪大学, 産業科学研究所, 助手 (10150658) 下村正二農林水産省, 農業生物資源研究所, 室長 (90116046) 前田正知大阪大学, 産業科学研究所, 助教授 (80190297)
Keywords	ATP / H^+ーATPase / H^+輸送 / 胃酸分泌 / H^+ / K^+ーATPase
Research Abstract	ATPの水解と共役してイオンを輸送するATPaseのうち、H^+ーATPaseとH^+/K^+ーATPaseのH^+輸送路、活性中心及びATP分解とH^+輸送の共役機構を明らかにすることを目的に研究を行なった。大腸菌H^+ーATPaseに変異を導入し、変異酵素の解析を通じ各アミノ酸残基の役割を推定した。明らかになった重要な点として、(1)βサブユニットのGlyーGlyーAlaーGlyーValーGlyーLysーThr(149ー156残基)がATPのγ位のリン酸基の近くにあること、Lysー155残基がLysー201残基に近いこと、Thrー156残基がMg^<2+>イオン及びSerー174残基に近いこと、Thrー156残基は本酵素の活性にとって必須であること。(2)本酵素のa__ーサブユニットがH^+の輸送路を形成しており、Argー210がおそらくH^+の授受に関与していること。(3)γサブユニットのカルボキシル末端がH^+輸送とATPの合成(分解)との共役の調節、おそらくゲ-ト機能に関与していること、等である。シナプス小胞と神経分泌顆粒に存在するH^+ーATPaseの構造と反応機構についても解析を行なった。このH^+ーATPaseの活性中心部分及びH^+輸送路ともに大腸菌H^+ーATPaseとよく似ていることが明らかになった。さらにこのATPaseの形成するH^+の電気化学的ポテンシャルが、神経伝達物質及びホルモンの輸送のエネルギ-となっていることを示した。このタイプのH^+ーATPaseのみが抗生物質バフィロマイシンによって阻害を受けることを見いだした点は、重要である。胃酸の分泌に関与するH^+/K^+ーATPaseについては胃壁細胞においてのみ遺伝子が転写されていること、この転写が胃壁細胞の核に存在するタンパクによって制御されていること、を明らかにした。ATPの類似化合物を用いる化学修飾によって本酵素のATP結合部位を明らかにした。このようにATPを分解しH^+を輸送するATPaseに関し、多くの新しい知見を得た。本研究によってATPの分解という化学反応とH^+の輸送というベクトル反応の共役機構を普遍的に理解できるようになろうとしている。

Research Products
(12 results)

All Other

All Publications (12 results)

[Publications] Atsuko Iwamoto: "H^+ーATPase γ subunit from <Escherichia>___ー <coli>___ー:Role of the conserved carboxyl terminal region." The Journal of Biological Chemistry. 265. 5043-5048 (1990)
[Publications] Masatomo Maeda: "Human gastric (H^++K^+)ーATPase gene: Similarity to (Na^++K^+)ーATPase genes in exon/intron organization but difference in control region." The Journal of Biological Chemistry. 265. 9027-9032 (1990)
[Publications] Yoshinori Moriyama: "Presence of 5'ーhydroxytryptamine (serotonine) transport coupled with vacuolar type H^+ーATPase in neurosecretory granules from bovine posterior pituitary." The Journal of Biological Chemistry. 265. 9165-9169 (1990)
[Publications] Hironori Hanada: "kinetic studies of chromaffin granule H^+ーATPase and effects of bafilomycin A_1." Biochemical and Biophysical Research Communications. 170. 873-878 (1990)
[Publications] Masaya Fujita: "Characterization of isoamylaseーhyperproducing mutant of <Pseudomonas>___ー <amyloderamosa>___ー." Agricultural and Biological Chemistry. 54. 2315-2321 (1990)
[Publications] Yoshinori Moriyama: "Energy coupling of Lーglutamate transport and vacuolar H^+ーATPase in brain synaptic vesicles." The Journal of Biochemistry. 108. 689-693 (1990)
[Publications] Michiyasu Takeyama: "The glycineーrich sequence of the β subunit of <Escherichia>___ー <coli>___ー H^+ーATPase is important for activity." The Journal of Biological Chemistry. 265. 21279-21284 (1990)
[Publications] Toshiyuki Umata: "The cytotoxic action of diphtheria toxin and its degradation in intact Vero celle are inhibited by bafilomycin Al,a specific inhibitor of vacuolarーtype H^+ーATPase." The Journal of Biological Chemistry. 265. 21940-21945 (1990)
[Publications] Yoshinori Moriyama: "H^+ーATPase,a primary pump for accumulation of neurotransmitters,is a major constituent of brain synaptic vesicls." Biochemical and Biophysical Researc Communications. 173. 443-448 (1990)
[Publications] Masamitsu Futai: "Molecular Structure,Function,and Assembly of the ATP Synthase" Plenum Press,New York, 9-18 (1990)
[Publications] Masamitsu Futai: "“Protein Engineering",Protein design in basic research,midicine,and industry" Japan Scientific Societies,and SpringerーVerlag, 173-180 (1990)
[Publications] Masatomo Maeda: "Bioenergetics: Molecular Biology,Biochemistry and Pathology" Plenum Press,New York, 217-225 (1990)

1990 Fiscal Year Annual Research Report

H^+ーATPaseおよびH^+/K^+ ATPaseによるイオン輸送機構の解析

Principal Investigator

二井將光大阪大学, 産業科学研究所, 教授 (50012646)

Research Products

[Publications] Atsuko Iwamoto: "H^+ーATPase γ subunit from <Escherichia>_ー <coli>_ー:Role of the conserved carboxyl terminal region." The Journal of Biological Chemistry. 265. 5043-5048 (1990)

[Publications] Masatomo Maeda: "Human gastric (H^++K^+)ーATPase gene: Similarity to (Na^++K^+)ーATPase genes in exon/intron organization but difference in control region." The Journal of Biological Chemistry. 265. 9027-9032 (1990)

[Publications] Yoshinori Moriyama: "Presence of 5'ーhydroxytryptamine (serotonine) transport coupled with vacuolar type H^+ーATPase in neurosecretory granules from bovine posterior pituitary." The Journal of Biological Chemistry. 265. 9165-9169 (1990)

[Publications] Hironori Hanada: "kinetic studies of chromaffin granule H^+ーATPase and effects of bafilomycin A_1." Biochemical and Biophysical Research Communications. 170. 873-878 (1990)

[Publications] Masaya Fujita: "Characterization of isoamylaseーhyperproducing mutant of <Pseudomonas>_ー <amyloderamosa>_ー." Agricultural and Biological Chemistry. 54. 2315-2321 (1990)

[Publications] Yoshinori Moriyama: "Energy coupling of Lーglutamate transport and vacuolar H^+ーATPase in brain synaptic vesicles." The Journal of Biochemistry. 108. 689-693 (1990)

[Publications] Michiyasu Takeyama: "The glycineーrich sequence of the β subunit of <Escherichia>_ー <coli>_ー H^+ーATPase is important for activity." The Journal of Biological Chemistry. 265. 21279-21284 (1990)

[Publications] Toshiyuki Umata: "The cytotoxic action of diphtheria toxin and its degradation in intact Vero celle are inhibited by bafilomycin Al,a specific inhibitor of vacuolarーtype H^+ーATPase." The Journal of Biological Chemistry. 265. 21940-21945 (1990)

[Publications] Yoshinori Moriyama: "H^+ーATPase,a primary pump for accumulation of neurotransmitters,is a major constituent of brain synaptic vesicls." Biochemical and Biophysical Researc Communications. 173. 443-448 (1990)

[Publications] Masamitsu Futai: "Molecular Structure,Function,and Assembly of the ATP Synthase" Plenum Press,New York, 9-18 (1990)

[Publications] Masamitsu Futai: "“Protein Engineering",Protein design in basic research,midicine,and industry" Japan Scientific Societies,and SpringerーVerlag, 173-180 (1990)

[Publications] Masatomo Maeda: "Bioenergetics: Molecular Biology,Biochemistry and Pathology" Plenum Press,New York, 217-225 (1990)

1990 Fiscal Year Annual Research Report

H^+ーATPaseおよびH^+/K^+ ATPaseによるイオン輸送機構の解析

Principal Investigator

二井 將光 大阪大学, 産業科学研究所, 教授 (50012646)

Research Products

[Publications] Atsuko Iwamoto: "H^+ーATPase γ subunit from <Escherichia>___ー <coli>___ー:Role of the conserved carboxyl terminal region." The Journal of Biological Chemistry. 265. 5043-5048 (1990)

[Publications] Masatomo Maeda: "Human gastric (H^++K^+)ーATPase gene: Similarity to (Na^++K^+)ーATPase genes in exon/intron organization but difference in control region." The Journal of Biological Chemistry. 265. 9027-9032 (1990)

[Publications] Yoshinori Moriyama: "Presence of 5'ーhydroxytryptamine (serotonine) transport coupled with vacuolar type H^+ーATPase in neurosecretory granules from bovine posterior pituitary." The Journal of Biological Chemistry. 265. 9165-9169 (1990)

[Publications] Hironori Hanada: "kinetic studies of chromaffin granule H^+ーATPase and effects of bafilomycin A_1." Biochemical and Biophysical Research Communications. 170. 873-878 (1990)

[Publications] Masaya Fujita: "Characterization of isoamylaseーhyperproducing mutant of <Pseudomonas>___ー <amyloderamosa>___ー." Agricultural and Biological Chemistry. 54. 2315-2321 (1990)

[Publications] Yoshinori Moriyama: "Energy coupling of Lーglutamate transport and vacuolar H^+ーATPase in brain synaptic vesicles." The Journal of Biochemistry. 108. 689-693 (1990)

[Publications] Michiyasu Takeyama: "The glycineーrich sequence of the β subunit of <Escherichia>___ー <coli>___ー H^+ーATPase is important for activity." The Journal of Biological Chemistry. 265. 21279-21284 (1990)

[Publications] Toshiyuki Umata: "The cytotoxic action of diphtheria toxin and its degradation in intact Vero celle are inhibited by bafilomycin Al,a specific inhibitor of vacuolarーtype H^+ーATPase." The Journal of Biological Chemistry. 265. 21940-21945 (1990)

[Publications] Yoshinori Moriyama: "H^+ーATPase,a primary pump for accumulation of neurotransmitters,is a major constituent of brain synaptic vesicls." Biochemical and Biophysical Researc Communications. 173. 443-448 (1990)

[Publications] Masamitsu Futai: "Molecular Structure,Function,and Assembly of the ATP Synthase" Plenum Press,New York, 9-18 (1990)

[Publications] Masamitsu Futai: "“Protein Engineering",Protein design in basic research,midicine,and industry" Japan Scientific Societies,and SpringerーVerlag, 173-180 (1990)

[Publications] Masatomo Maeda: "Bioenergetics: Molecular Biology,Biochemistry and Pathology" Plenum Press,New York, 217-225 (1990)

二井將光大阪大学, 産業科学研究所, 教授 (50012646)

[Publications] Atsuko Iwamoto: "H^+ーATPase γ subunit from <Escherichia>_ー <coli>_ー:Role of the conserved carboxyl terminal region." The Journal of Biological Chemistry. 265. 5043-5048 (1990)

[Publications] Masaya Fujita: "Characterization of isoamylaseーhyperproducing mutant of <Pseudomonas>_ー <amyloderamosa>_ー." Agricultural and Biological Chemistry. 54. 2315-2321 (1990)

[Publications] Michiyasu Takeyama: "The glycineーrich sequence of the β subunit of <Escherichia>_ー <coli>_ー H^+ーATPase is important for activity." The Journal of Biological Chemistry. 265. 21279-21284 (1990)