1989 年度実績報告書

低分子量GTP結合蛋白質を介する新しい細胞内情報伝達機構

研究課題

研究課題/領域番号	01480529
研究機関	神戸大学
研究代表者	高井義美神戸大学, 医学部, 教授 (60093514)
研究分担者	溝口明神戸大学, 医学部, 講師 (90181916) 菊池章神戸大学, 医学部, 助手 (10204827) 貝淵弘三神戸大学, 医学部, 講師 (00169377)
キーワード	GTP結合蛋白質 / smgp21 / smgp25A / GAP / GDI
研究概要	私共は分子量2万余りの低分子量GTP結合蛋白質(G蛋白質)をウシ大脳から15種類以上分離し、そのうち5種類(CーKi-rasp21、rhoAp21、rhoBp20、smgp21、smgp25A)を精製している。これらの低分子量G蛋白質にはGDP結合型の不活性型とGTP結合型の活性型が存在し、不活性型と活性型の転換を制御する蛋白質や活性型が作用する標的蛋白質が存在すると考えられている。本研究では、低分子量G蛋白質の活性制御蛋白質の精製をまず試み、smgp21とrhoBp20のGTPase活性を促進する蛋白質(smgp21GAPs、rhoGAP)を精製した。また、smgp25AのGDPの解離を抑制し、その結果GTPの結合を抑制する全く新しい種類の蛋白質、GDP解離抑制蛋白質(smgp25AGDI)も精製することに成功した。さらに、rhoBp20に対するGDI(rhoGDI)も精製し、smgp25AGDIとrhoGDIのCDNAを単離して一次構造を決定した。以上の結果から、低分子量G蛋白質の活性はGAPとGDIの少なくとも2種類の活性制御蛋白質により制御されていることが明らかとなった。次に、smgp21とsmgp25Aに対する特異抗体を作製し、その組織内分布と細胞内分布を検討した。smgp21はrasp21と同じエフェクタ-部位をもつことから、rasp21と同じかあるいは拮抗する作用を有すると考えられる。しかし、ある種の組織や細胞では、smgp21はrasp21とは異なる組織分布や細胞内分布を示し、smgp21はrasp21とは異なるそれ固有の作用を有していることが示唆された。また、smgp25Aは神経細胞や分泌細胞に存在し、特に脳内ではシナプスに大量に存在することから、smgp25Aが分泌反応に関与している可能性が示唆された。今後は、上述した2種類の活性制御蛋白質のCDNAや特異抗体を用いて、低分子量G蛋白質を介する新しい細胞内情報伝達系の生理的な役割を解明していく予定である。以上、本年度の研究計画はほぼ達成することができた。

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] Ohmori,T.: "Small molecular weight GTP-binding proteins in human platelet membranes--Purification and characterization of a novel GTP-binding protein with a molecular weight of 22,000." J.Biol.Chem.264. 1877-1881 (1989)
[文献書誌] Kikuchi,A.: "Purification and characterization from bovine brain cytosol of two GTPase-activating proteins specific for smg p21,a GTP-binding protein having the same effector domain as c-ras p21s." J.Biol.Chem.264. 9133-9136 (1989)
[文献書誌] Kaibuchi,K.: "Molecular genetic analysis of the regulatory and catalytic domains of protein kinase C." J.Biol.Chem.264. 13489-13496 (1989)
[文献書誌] Kawata,M.: "Phosphorylation of smg p21,a ras p21-like GTP-binding protein,by cyclic AMP-dependent protein kinase in a cell-free system and in response to prostaglandin E_1 in intact human platelets." J.Biol.Chem.264. 15688-15695 (1989)
[文献書誌] Kaibuchi,K.: "Activation of the serum-response element and TPA-response element by the activated c-raf-1 protein in a manner independent of protein kinase C." J.Biol.Chem.264. 20855-20858 (1989)
[文献書誌] Mizoguchi,A.: "Localization and subcellular distribution of cellular ras gene products in rat brain." Mol.Brain Res.5. 31-44 (1989)
[文献書誌] Kim,S.: "Intrasynaptosomal distribution of the ras,rho and smg-25A GTP-binding proteins in bovine brain." Mol.Brain Res.6. 167-176 (1989)
[文献書誌] Sano,K.: "Tissue-specific expression of a novel GTP-binding protein(smg p25A)mRNA and its increase by nerve growth factor and cyclic AMP in rat pheochromocytoma PC-12 cells." Biochem.Biophys.Res.Commun.158. 377-385 (1989)
[文献書誌] Ueda,T.: "GTPase activating proteins for the smg-21 GTP-binding protein having the same effector domain as the ras proteins in human platelets." Biochem.Biophys.Res.Commun.159. 1411-1419 (1989)
[文献書誌] Shoji,I.: "Kinetic analysis of the binding of guanine nucleotide to bovine brain smg p25A." Biochem.Biophys.Res.Commun.162. 273-281 (1989)
[文献書誌] Mizoguchi,A.: "Tissue distribution of smg p25A,A ras p21-like GTP-binding protein,studied by use of a specific monoclonal antibody." Biochem.Biophys.Res.Commun.162. 1438-1445 (1989)
[文献書誌] Kuroda,S.: "Kinetic analysis of the binding of guanine nucleotides to bovine brain rhoB p20,a ras p21-like GTP-binding protein." Biochem.Biophys.Res.Commun.163. 674-681 (1989)
[文献書誌] Kawata,M.: "Identification of a major GTP-binding protein in bovine aortic smooth muscle membranes as smg p21,a GTP-binding protein having the same effector domain as ras p21s." Biochem.Biophys.Res.Commun.163. 1418-1427 (1989)
[文献書誌] Ohga,N.: "Rabbit intestine contains a protein that inhibits the dissociation of GDP from and the subsequent binding of GTP to rhoB p20,a ras p21-like GTP-binding protein." Biochem.Biophys.Res.Commun.163. 1523-1533 (1989)
[文献書誌] Yamamoto,K.: "Small molecular weight GTP-binding proteins and signal transduction." Clinica.Chimica.Acta.185. 347-356 (1989)
[文献書誌] Fukumoto,Y.: "Activation of the c-fos serum-response element by the activated c-Ha-ras protein in a manner independent of protein kinase C and cAMP-dependent protein kinase." J.Biol.Chem.265. 774-780 (1990)
[文献書誌] Sasaki,T.: "Purification and characterization from bovine brain cytosol of a protein that inhibits the dissociation of GDP from and the subsequent binding of GTP to smg p25A,a ras p21-like GTP-binding protein." J.Biol.Chem.265. 2333-2337 (1990)
[文献書誌] Hoshijima,M.: "Purification and characterization from bovine brain membranes of a GTP-binding protein with a Mr of 21,000 ADP-ribosylated by an ADP-ribosyltransferase contained in botulinum toxin type C1--Identification as the rhoA gene product." Mol.Brain Res.7. 9-16 (1990)
[文献書誌] Matsui,Y.: "Molecular cloning of smg p21B and identification of smg p21 purified from bovine brain and human platelets as smg p21B." Biochem.Biophys.Res.Commun.166. 1010-1016 (1990)
[文献書誌] Hata,Y.: "Inhibition of the ras p21 GTPase-activating protein-stimulated GTPase activity of c-Ha-ras p21 by smg p21 having the same putative effector domain as ras p21s." J.Biol.Chem.(1990)
[文献書誌] Yamamoto,J.: "A GTPase-activating protein(GAP)for rhoB p20,a ras p21-like GTP-binding protein--Partial purification,characterization and subcellular distribution in rat brain." Mol.Brain Res.(1990)
[文献書誌] Kim,S.: "Tissue and subcellular distributions of smg p21s,ras p21-like GTP-binding proteins having the same putative effector domain as ras p21s,partly distinct from those of c-ras p21s." Mol.Cell.Biol.(1990)

1989 年度 実績報告書

低分子量GTP結合蛋白質を介する新しい細胞内情報伝達機構

研究代表者

高井 義美 神戸大学, 医学部, 教授 (60093514)

研究成果

[文献書誌] Ohmori,T.: "Small molecular weight GTP-binding proteins in human platelet membranes--Purification and characterization of a novel GTP-binding protein with a molecular weight of 22,000." J.Biol.Chem.264. 1877-1881 (1989)

[文献書誌] Kikuchi,A.: "Purification and characterization from bovine brain cytosol of two GTPase-activating proteins specific for smg p21,a GTP-binding protein having the same effector domain as c-ras p21s." J.Biol.Chem.264. 9133-9136 (1989)

[文献書誌] Kaibuchi,K.: "Molecular genetic analysis of the regulatory and catalytic domains of protein kinase C." J.Biol.Chem.264. 13489-13496 (1989)

[文献書誌] Kawata,M.: "Phosphorylation of smg p21,a ras p21-like GTP-binding protein,by cyclic AMP-dependent protein kinase in a cell-free system and in response to prostaglandin E_1 in intact human platelets." J.Biol.Chem.264. 15688-15695 (1989)

[文献書誌] Kaibuchi,K.: "Activation of the serum-response element and TPA-response element by the activated c-raf-1 protein in a manner independent of protein kinase C." J.Biol.Chem.264. 20855-20858 (1989)

[文献書誌] Mizoguchi,A.: "Localization and subcellular distribution of cellular ras gene products in rat brain." Mol.Brain Res.5. 31-44 (1989)

[文献書誌] Kim,S.: "Intrasynaptosomal distribution of the ras,rho and smg-25A GTP-binding proteins in bovine brain." Mol.Brain Res.6. 167-176 (1989)

[文献書誌] Sano,K.: "Tissue-specific expression of a novel GTP-binding protein(smg p25A)mRNA and its increase by nerve growth factor and cyclic AMP in rat pheochromocytoma PC-12 cells." Biochem.Biophys.Res.Commun.158. 377-385 (1989)

[文献書誌] Ueda,T.: "GTPase activating proteins for the smg-21 GTP-binding protein having the same effector domain as the ras proteins in human platelets." Biochem.Biophys.Res.Commun.159. 1411-1419 (1989)

[文献書誌] Shoji,I.: "Kinetic analysis of the binding of guanine nucleotide to bovine brain smg p25A." Biochem.Biophys.Res.Commun.162. 273-281 (1989)

[文献書誌] Mizoguchi,A.: "Tissue distribution of smg p25A,A ras p21-like GTP-binding protein,studied by use of a specific monoclonal antibody." Biochem.Biophys.Res.Commun.162. 1438-1445 (1989)

[文献書誌] Kuroda,S.: "Kinetic analysis of the binding of guanine nucleotides to bovine brain rhoB p20,a ras p21-like GTP-binding protein." Biochem.Biophys.Res.Commun.163. 674-681 (1989)

[文献書誌] Kawata,M.: "Identification of a major GTP-binding protein in bovine aortic smooth muscle membranes as smg p21,a GTP-binding protein having the same effector domain as ras p21s." Biochem.Biophys.Res.Commun.163. 1418-1427 (1989)

[文献書誌] Ohga,N.: "Rabbit intestine contains a protein that inhibits the dissociation of GDP from and the subsequent binding of GTP to rhoB p20,a ras p21-like GTP-binding protein." Biochem.Biophys.Res.Commun.163. 1523-1533 (1989)

[文献書誌] Yamamoto,K.: "Small molecular weight GTP-binding proteins and signal transduction." Clinica.Chimica.Acta.185. 347-356 (1989)

[文献書誌] Fukumoto,Y.: "Activation of the c-fos serum-response element by the activated c-Ha-ras protein in a manner independent of protein kinase C and cAMP-dependent protein kinase." J.Biol.Chem.265. 774-780 (1990)

[文献書誌] Sasaki,T.: "Purification and characterization from bovine brain cytosol of a protein that inhibits the dissociation of GDP from and the subsequent binding of GTP to smg p25A,a ras p21-like GTP-binding protein." J.Biol.Chem.265. 2333-2337 (1990)

[文献書誌] Hoshijima,M.: "Purification and characterization from bovine brain membranes of a GTP-binding protein with a Mr of 21,000 ADP-ribosylated by an ADP-ribosyltransferase contained in botulinum toxin type C1--Identification as the rhoA gene product." Mol.Brain Res.7. 9-16 (1990)

[文献書誌] Matsui,Y.: "Molecular cloning of smg p21B and identification of smg p21 purified from bovine brain and human platelets as smg p21B." Biochem.Biophys.Res.Commun.166. 1010-1016 (1990)

[文献書誌] Hata,Y.: "Inhibition of the ras p21 GTPase-activating protein-stimulated GTPase activity of c-Ha-ras p21 by smg p21 having the same putative effector domain as ras p21s." J.Biol.Chem.(1990)

[文献書誌] Yamamoto,J.: "A GTPase-activating protein(GAP)for rhoB p20,a ras p21-like GTP-binding protein--Partial purification,characterization and subcellular distribution in rat brain." Mol.Brain Res.(1990)

[文献書誌] Kim,S.: "Tissue and subcellular distributions of smg p21s,ras p21-like GTP-binding proteins having the same putative effector domain as ras p21s,partly distinct from those of c-ras p21s." Mol.Cell.Biol.(1990)

1989 年度実績報告書

高井義美神戸大学, 医学部, 教授 (60093514)