1993 Fiscal Year Annual Research Report

イオン・溶質輸送と共役ネットワーク

Research Project

Project/Area Number	04266101
Research Institution	Tokyo Metropolitan Organization for Medical Research
Principal Investigator	川喜田正夫財団法人東京都臨床医学総合研究所, 医科学研究部門, 研究員 (00012740)
Co-Investigator(Kenkyū-buntansha)	原諭吉東京医科歯科大学, 医学部, 助教授 (00092429) 山口明人千葉大学, 薬学部, 助教授 (60114336) 重川宗一国立循環器センター研究所, 循環器分子生理部, 研究員 (00113738) 青島均山口大学, 教養部, 教授 (20089907) 乾賢一東京医歯大, 医学部, 教授 (70034030)
Keywords	イオン輸送 / 共役物質輸送 / Na^+ / H^+-アンチポーター / ジペプチド輸送系 / H^+ヘキソース共輸送 / Na^+ / K^+-ATPase / テトラサイクリン排出タンパク質
Research Abstract	細胞の膜系における各種のイオンや溶質の共役輸送系の実態とそのネットワークに関して、以下の結果を得た。 1.ウシ腎臓及び小腸の刷子縁膜に存在するNa^+/H^+-アンチポーターの分子種が、抗体との反応性、阻害剤感受性、反応速度論的性質等の観点からみて、異なる分子種であることを示した(川喜田)。 2.Na^+/H^+-アンチポーターのNHE1 isoformが高親和性カルモジュリン(CaM)結合タンパク質であることを明らかにし、増殖因子によるNHE1の活性化において、CaMとの相互作用が重要な役割を果たす可能性が強いことを示した(重川)。 3.食細胞のH^+サーキットの活性化に関連して、NADPHオキシダーゼ活性化の分子機構を解析し、p47タンパク質SH2領域の重要性を示す結果を得た(水上)。 4.多孔性フィルター上に培養した上皮細胞シートによる物質輸送のin vitro評価系を確立し、腸管のジペプチド経細胞輸送に頂端膜側ではH^+共役型、側底膜側ではH^+非依存型輸送系が関与することを確立した(乾)。 5.アフリカツメガエル卵母細胞にH^+/ヘキソース共輸送体mRNAを注入し、基質の添加に伴う輸送電流を測定する方法を確立し、各種の糖に対する速度論的パラメーターを測定することに成功した(青島)。 6.Na^+/K^+-ATPaseをBALB/c3T3細胞中で発現させ、Naポンプ電流を測定する方法を確立し、K^+の膜内移動過程の解析から輸送過程におけるK^+の荷電状態を明らかにした(原)。 7.テトラサイクリン排出タンパク質の部位特異的変異体約300種類を作製し、その解析を通じて輸送機能に必須な残基の分布とそれらの役割を明らかにした。さらに、特異的修飾試薬に対する反応性の解析とあわせて、輸送モデル、gated single-pathwayモデルを提案した(山口)。

Research Products
(36 results)

All Other

All Publications (36 results)

[Publications] Yamashita,T.: "Characterization of the Na^+/H^+-antiporter of bovine renal cortex and its immunoaffiruity purification in a reconstitutively active form" J.Biochem.113. 136-140 (1993)
[Publications] Suzuki,T.: "Uncompling of ATP splitting from Ca^<2+>-transport in Ca^<2+>-transporting ATPase of the Sarcoplasmic Reticulum as a result of modification by N-(3-pyrene) maleimide:activation of a channel with a specificity for alkaline earth metal ions" J.Biochem.114. 203-209 (1993)
[Publications] Kuroda,M.: "Calcium pump expresion in human bone and soft tissue tumors" Virehows Arehiv A Pathol.Anat.423. 209-213 (1993)
[Publications] Hara,T.: "The UDP-galactosetranslocator gene is mapped to the Wiscott-Aldrich syndrome locus of the X chromosome" Somat.Cell Mol.Genet. (in press).
[Publications] Yamamoto,K.: "Regulation of Na,K-adenosine triphospjatase gene sxpression by sodium ioms in cultured neonatal rat cardiocytes" J.Clinical Investigations. 92. 1889-1895 (1993)
[Publications] Aoshima,H.: "Heterologans expression of the H^+/hexose cotransporter from chrorella in xenopus oocytes and its characterization" J.Plant Physiol.141. 293-297 (1993)
[Publications] Sumimoto,H: "Role of Src homology 3(SH3)domains in assembly and activation of the phagacyte NADPH oxidase" Proc.Natl.Acad.Sci.USA. (in press).
[Publications] Sumimoto,H: "Formation of a nonvel 20-hydroxylated metablite of lipoxin A_4 by human neutrophil microsomes" FEBS Lett.315. 205-210 (1993)
[Publications] Mizukami,Y: "w-Hydroxylation of lipoxin B_4 by human neutrophil microsomes:identification of w-hydroxy metabolite of lipoxin B_4 and catalysis by leukotriene B_4 w-hydroxylase (cytochrome P-450 LTBw)" Biochim.Biophys.Acta.1168. 87-93 (1993)
[Publications] Yu,L: "Characterization of the GTP-dependent activation of the superoxide-producing NADPH oxidase in cell-free system of pig neutrophils" Biochim.Biophys.Acta.1178. 73-80 (1993)
[Publications] Mitsuyama,T: "Cyclic AMP inhibits the respiratory burst of electropermeabilized human neutrophils at a downstream site of protein kinase C" Biochim.Biophys.Acta.1177. 167-173 (1993)
[Publications] Kansha,M: "Decrease in the phosphotyrosine phosphatase activity in the plasma membrane of human neutrophils on stimulation by phorbol 12-myristate 13-acetate" Biochim.Biophys.Acta.1179. 189-196 (1993)
[Publications] H.Saito: "Dipeptide transporters in apical and basolateral membranes of the human intestinal cell line CaCo-2" Am.J.Physiol.265. G289-G294 (1993)
[Publications] H.Saito: "Expression of human intestinal dipeptide transporter in xenopus laevis occytes" Biochem.Pharmacol. 45. 776-779 (1993)
[Publications] R.Hori: "Transport of bestatin in rat renal brush-border membrane vesicles" Biochem.Pharmacol.45. 1763-1768 (1993)
[Publications] K.Ohoka: "p-Aminohippurate transport in rat renal brush-border membranes:a potential-sensitive transport system and an anion exchanger" Biol.Pharm.Bull.16. 395-401 (1993)
[Publications] T.Katsura: "Characteristics of organic cation transporter in rat renal basolateral membrane" Biochim.Biophys.Acta. 1146. 197-202 (1993)
[Publications] S.Maeda: "Transport of organic cation in renal brush-border membrane from rats with renal ischemic injury" Biochim.Biophys.Acta. 1150. 103-110 (1993)
[Publications] Bertrand,B.et al.: "The Na^+/H^+ exchanger isoform 1 (NHE1) is a novel member of calmodulin binding proteins." J.Biol.Chem.(in press). (1994)
[Publications] Wakabayashi,S.et al.: "Mutaion of calmodulin-binding site renders the Na^+/H^+ exchanger (NHE1) highly H^+-sensitive and Ca^<2+> regulation-defective." J.Biol.Chem.(in press). (1994)
[Publications] Wakabayashi,S.et al.: "Growth factor activation and "H^+-sensing" of the Na^+/H^+ exchanger isoform 1 (NHE1).Evidence for an additional mechanism not requirin direct phosphoryla" J.Biol.Chem.(in press). (1994)
[Publications] Nakasaki,Y.et al.: "Cloning of the rat aortic smooth muscle Na/Ca exchanger and tissue-specific expression of isoforms." J.Biochem.114. 528-534 (1993)
[Publications] Taniguchi,J.et al.: "Maxi K^+ channels are stimulated by cyclic guanosine monophosphate-dependent protein kinase in canine coronary artery smooth muscle cell" Pflugers Arch.423. 167-172 (1993)
[Publications] Tawada-Iwata,Y.et al.: "Invreased mechanical extraction of T-tuble/junctional SR from cardiomyopathic hamster heart." Am.J.Physiol.264. H1447-H1453 (1993)
[Publications] Yamaguchi,A.et al.: "Metal-Tetracycline/H^+ Antiporter of Escherichia coli Encoded by Transposon Tn10:The Structural......" J.Biol.Chem.268. 6496-6504 (1993)
[Publications] Yamaguchi,A.et al.: "Role of the Conserved Quartet of Residues Located in the N- and C-Terminal Halves of the Transposon Tn10.." Biochemistry. 32. 5698-5704 (1993)
[Publications] Yamaguchi,A.et al.: "The in vivo Assembly of Function of the N- and C-Ter-minal Halves of the Tn10-Encoded TetA protein" FEBS Letters. 324. 131-135 (1993)
[Publications] Yamaguchi,A.et al.: "Effects of Sulfhydryl Reagents on the Cys65 Mutant of the Trans-poson Tn10-Encoded Metal-Tetracyclin/H^+ Antiporter.of E.coli" FEBS Letters. 322. 201-204 (1993)
[Publications] Yamaguchi,A.et al.: "Second-Site Mutation of Ala-220 to Glu or Asp Suppresses the Mutation of Asp-285 to Asn in the Transposon Tn10-Encoded Metal.." J.Biol.Chem.268. 26990-26995 (1993)
[Publications] Yamaguchi,A.et al.: "Hot-Spots for Sulfhydryl Inactivation of Cys Mutants in the Widely Conserved Sequence Motifs of the Metal-Tetracycline/H^+..." J.Biochemistry. 115(in press). (1994)
[Publications] Hara,Y.: "Kinetic analysis and voltage dependency of alpha-3 Na^+/K^+-pump in proceedimgs for the 7th international conference on Na pump" Dr.D Steinkopff Verlag,Darmstadt (in print),
[Publications] 青島均: 朝倉書店 (井村裕夫,岡哲雄,芳賀達也,岩本英爾編)27HC02:レセプター -基礎と臨床中.ニコチン性アセチルコリンレセプター, 944 pp.400-405 (1993)
[Publications] Takeshige,K: "Active Oxygens,Lipid Peroxides,and Antioxidants" Japan Sci.Soc.Press, 13 (1993)
[Publications] 乾賢一: "膜学実験シリーズ第I 巻生体膜編日本膜学会編" 共立出版, 424 (1994)
[Publications] 橋本一,井上松久編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)
[Publications] 橋本一,井上松久編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)(994)

1993 Fiscal Year Annual Research Report

イオン・溶質輸送と共役ネットワーク

Principal Investigator

川喜田 正夫 財団法人 東京都臨床医学総合研究所, 医科学研究部門, 研究員 (00012740)

Research Products

[Publications] Yamashita,T.: "Characterization of the Na^+/H^+-antiporter of bovine renal cortex and its immunoaffiruity purification in a reconstitutively active form" J.Biochem.113. 136-140 (1993)

[Publications] Suzuki,T.: "Uncompling of ATP splitting from Ca^<2+>-transport in Ca^<2+>-transporting ATPase of the Sarcoplasmic Reticulum as a result of modification by N-(3-pyrene) maleimide:activation of a channel with a specificity for alkaline earth metal ions" J.Biochem.114. 203-209 (1993)

[Publications] Kuroda,M.: "Calcium pump expresion in human bone and soft tissue tumors" Virehows Arehiv A Pathol.Anat.423. 209-213 (1993)

[Publications] Hara,T.: "The UDP-galactosetranslocator gene is mapped to the Wiscott-Aldrich syndrome locus of the X chromosome" Somat.Cell Mol.Genet. (in press).

[Publications] Yamamoto,K.: "Regulation of Na,K-adenosine triphospjatase gene sxpression by sodium ioms in cultured neonatal rat cardiocytes" J.Clinical Investigations. 92. 1889-1895 (1993)

[Publications] Aoshima,H.: "Heterologans expression of the H^+/hexose cotransporter from chrorella in xenopus oocytes and its characterization" J.Plant Physiol.141. 293-297 (1993)

[Publications] Sumimoto,H: "Role of Src homology 3(SH3)domains in assembly and activation of the phagacyte NADPH oxidase" Proc.Natl.Acad.Sci.USA. (in press).

[Publications] Sumimoto,H: "Formation of a nonvel 20-hydroxylated metablite of lipoxin A_4 by human neutrophil microsomes" FEBS Lett.315. 205-210 (1993)

[Publications] Mizukami,Y: "w-Hydroxylation of lipoxin B_4 by human neutrophil microsomes:identification of w-hydroxy metabolite of lipoxin B_4 and catalysis by leukotriene B_4 w-hydroxylase (cytochrome P-450 LTBw)" Biochim.Biophys.Acta.1168. 87-93 (1993)

[Publications] Yu,L: "Characterization of the GTP-dependent activation of the superoxide-producing NADPH oxidase in cell-free system of pig neutrophils" Biochim.Biophys.Acta.1178. 73-80 (1993)

[Publications] Mitsuyama,T: "Cyclic AMP inhibits the respiratory burst of electropermeabilized human neutrophils at a downstream site of protein kinase C" Biochim.Biophys.Acta.1177. 167-173 (1993)

[Publications] Kansha,M: "Decrease in the phosphotyrosine phosphatase activity in the plasma membrane of human neutrophils on stimulation by phorbol 12-myristate 13-acetate" Biochim.Biophys.Acta.1179. 189-196 (1993)

[Publications] H.Saito: "Dipeptide transporters in apical and basolateral membranes of the human intestinal cell line CaCo-2" Am.J.Physiol.265. G289-G294 (1993)

[Publications] H.Saito: "Expression of human intestinal dipeptide transporter in xenopus laevis occytes" Biochem.Pharmacol. 45. 776-779 (1993)

[Publications] R.Hori: "Transport of bestatin in rat renal brush-border membrane vesicles" Biochem.Pharmacol.45. 1763-1768 (1993)

[Publications] K.Ohoka: "p-Aminohippurate transport in rat renal brush-border membranes:a potential-sensitive transport system and an anion exchanger" Biol.Pharm.Bull.16. 395-401 (1993)

[Publications] T.Katsura: "Characteristics of organic cation transporter in rat renal basolateral membrane" Biochim.Biophys.Acta. 1146. 197-202 (1993)

[Publications] S.Maeda: "Transport of organic cation in renal brush-border membrane from rats with renal ischemic injury" Biochim.Biophys.Acta. 1150. 103-110 (1993)

[Publications] Bertrand,B.et al.: "The Na^+/H^+ exchanger isoform 1 (NHE1) is a novel member of calmodulin binding proteins." J.Biol.Chem.(in press). (1994)

[Publications] Wakabayashi,S.et al.: "Mutaion of calmodulin-binding site renders the Na^+/H^+ exchanger (NHE1) highly H^+-sensitive and Ca^<2+> regulation-defective." J.Biol.Chem.(in press). (1994)

[Publications] Wakabayashi,S.et al.: "Growth factor activation and "H^+-sensing" of the Na^+/H^+ exchanger isoform 1 (NHE1).Evidence for an additional mechanism not requirin direct phosphoryla" J.Biol.Chem.(in press). (1994)

[Publications] Nakasaki,Y.et al.: "Cloning of the rat aortic smooth muscle Na/Ca exchanger and tissue-specific expression of isoforms." J.Biochem.114. 528-534 (1993)

[Publications] Taniguchi,J.et al.: "Maxi K^+ channels are stimulated by cyclic guanosine monophosphate-dependent protein kinase in canine coronary artery smooth muscle cell" Pflugers Arch.423. 167-172 (1993)

[Publications] Tawada-Iwata,Y.et al.: "Invreased mechanical extraction of T-tuble/junctional SR from cardiomyopathic hamster heart." Am.J.Physiol.264. H1447-H1453 (1993)

[Publications] Yamaguchi,A.et al.: "Metal-Tetracycline/H^+ Antiporter of Escherichia coli Encoded by Transposon Tn10:The Structural......" J.Biol.Chem.268. 6496-6504 (1993)

[Publications] Yamaguchi,A.et al.: "Role of the Conserved Quartet of Residues Located in the N- and C-Terminal Halves of the Transposon Tn10.." Biochemistry. 32. 5698-5704 (1993)

[Publications] Yamaguchi,A.et al.: "The in vivo Assembly of Function of the N- and C-Ter-minal Halves of the Tn10-Encoded TetA protein" FEBS Letters. 324. 131-135 (1993)

[Publications] Yamaguchi,A.et al.: "Effects of Sulfhydryl Reagents on the Cys65 Mutant of the Trans-poson Tn10-Encoded Metal-Tetracyclin/H^+ Antiporter.of E.coli" FEBS Letters. 322. 201-204 (1993)

[Publications] Yamaguchi,A.et al.: "Second-Site Mutation of Ala-220 to Glu or Asp Suppresses the Mutation of Asp-285 to Asn in the Transposon Tn10-Encoded Metal.." J.Biol.Chem.268. 26990-26995 (1993)

[Publications] Yamaguchi,A.et al.: "Hot-Spots for Sulfhydryl Inactivation of Cys Mutants in the Widely Conserved Sequence Motifs of the Metal-Tetracycline/H^+..." J.Biochemistry. 115(in press). (1994)

[Publications] Hara,Y.: "Kinetic analysis and voltage dependency of alpha-3 Na^+/K^+-pump in proceedimgs for the 7th international conference on Na pump" Dr.D Steinkopff Verlag,Darmstadt (in print),

[Publications] 青島 均: 朝倉書店 (井村 裕夫,岡 哲雄,芳賀 達也,岩本 英爾 編)27HC02:レセプター -基礎と臨床 中.ニコチン性アセチルコリンレセプター, 944 pp.400-405 (1993)

[Publications] Takeshige,K: "Active Oxygens,Lipid Peroxides,and Antioxidants" Japan Sci.Soc.Press, 13 (1993)

[Publications] 乾 賢一: "膜学実験シリーズ 第I 巻 生体膜編 日本膜学会編" 共立出版, 424 (1994)

[Publications] 橋本 一,井上 松久 編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)

[Publications] 橋本 一,井上 松久 編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)(994)

川喜田正夫財団法人東京都臨床医学総合研究所, 医科学研究部門, 研究員 (00012740)

[Publications] 青島均: 朝倉書店 (井村裕夫,岡哲雄,芳賀達也,岩本英爾編)27HC02:レセプター -基礎と臨床中.ニコチン性アセチルコリンレセプター, 944 pp.400-405 (1993)

[Publications] 乾賢一: "膜学実験シリーズ第I 巻生体膜編日本膜学会編" 共立出版, 424 (1994)

[Publications] 橋本一,井上松久編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)

[Publications] 橋本一,井上松久編: "病原菌の薬剤耐性機構の解明とその対策(共著)" 学会出版センター, 198 (1993)(994)