1992 Fiscal Year Annual Research Report

大腸菌蛋白質分泌の分子機構

Research Project

Project/Area Number	04259102
Research Institution	The University of Tokyo
Principal Investigator	水島昭二東京大学, 応用微生物研究所, 教授 (50013313)
Co-Investigator(Kenkyū-buntansha)	伊藤維昭京都大学, ウィルス研究所, 教授 (90027334)
Keywords	蛋白質の分泌 / 蛋白質の膜透過 / Sec蛋白質 / SecY / SecE / シグナルペプチド / 変異解析 / SecA
Research Abstract	I.生化学的研究(主として水島グループ) 1.SecEのドメイン機能の解析:SecEのC末端部分のうち、膜貫通領域がSecE機能の重要な部分を担っていることを明らかにした。 2.SecYの機能:SecYを大量精製する方法を確立し、SecYのX線結晶解析へ一歩前進した。またSecYがタンパク質の膜透過にともなうプロトンの対向輪送に重要な役割を果たしていることを示した。 3.SecD,SecFの機能解析:SecDがタンパク質の膜透過後の放出に重要であること、またSecFはSecDアッセンブリーに関与していることを示した。 4.シグナルペプチドN末正荷電領域の機能についての新知見を得た。また非ペプチド結合部分をもつタンパク質も膜透過しうることを示した。 II.遺伝学的研究(伊藤グループ) SecYの優性欠損変異を解析し、C末端側にSecY活性に重要な部位があることを提唱、その効果を打ち消す遺伝子内抑制変異の分離・解析から他の因子との相互作用に関わる部位がSecYの中央部位に存在する事を示唆した。この領域の変異secY24はSecEとの相互作用を弱める事を示した。SecD,SecF,および″ORF12″のsecYとの相互作用を調べ、ORF12がSecY安定化効果および優性欠損secY変異に対する抑制効果に関わることを見い出した。新しい遺伝子ydrの産物がSecYと相互作用を持つ事を示唆した。膜蛋白質の膜へのアンカリング効率が低下した変異をftsH遺伝子内に同定し、ATPase結合合部位の重要性、蛋白質膜透過自体への関与等を明らかにした。

Research Products

(16 results)

All Other

All Publications (16 results)

[Publications] M.Kato et al.: "In Vitro translocation of secretory proteins possessing no charges at the mature domain takes place efficiently in a protonmotive force-dependent manner." J.Biol.Chem.267. 413-418 (1992)
[Publications] L.Bruncage et al.: "SecY,SecE and band 1 form the membrane-embedded domain of E.coli preprotrin translocase." J.Biol Chem.267. 4166-4170 (1992)
[Publications] C.Hikita et al.: "Effects of total-hydrophobicity and length of the hydrophobic domain of a signal peptied on in vitro translocation effciency." J.Biol Chem.267. 4882-4888 (1992)
[Publications] K.Nishiyama et al.: "The C-terminal region of SecE interacts with SecY and id functional in the reconstitution of protein translocation activity in Escherichia coli." J.Biol Chem.267. 7170-7176 (1992)
[Publications] S.Matsuyama et al.: "Overproduction,purification and characterization of SecD and SecF,integral membrane components of the protein translocation machinery of Escherichia coli." Biochim.Biophys.Acta. 1122. 77-84 (1992)
[Publications] C.Hikita et al.: "The requirement of a positive charge at the amino terminus can be compensated for by a longer central hydrophobic stretch in the functioning of signala peptides." J.Biol.Chem.267. 12375-12379 (1992)
[Publications] H.Takamatsu et al.: "In vivo and in vitro characterization of the SecA gene product of bacillus subtilis." J.Bacteriol.174. 4308-4316 (1992)
[Publications] S,Matsuyama et al.: "Large-scale Production of membrane proteins fused to a truncated SecA in Escherichia coli." Biosci.Biotech.Biochem.56. 1512-1514 (1992)
[Publications] S.Kamitani te al.: "Identification and characterization of an Escherichia coli gene required for the formation of correctly folded alkaline phosphatase,a periplasmic enzyme." BMBO J.11. 57-62 (1992)
[Publications] C.Ueguchi et al.: "Multicopy suppression:an approach to understanding intracellular functioning of the protein export system." J.Bacteriol.174. 1454-1461 (1992)
[Publications] T.Shimoike et al.: "SecY variants that interfere with Escherichia coll protein export in the presence of normal secY." Mol.Microbiol.6. 1205-1210 (1992)
[Publications] K,Ito et al.: "SecY and integral membrane components of the Escherichia colf protein translocation ststem." Mol.Microbiol.6. 2423-2428 (1992)
[Publications] T.Taura et al.: "Insertional disruption of the nusB gene leads to cold-sensitive growth and suppression of the secY24 mutation." Mol.Gen.Genet.234. 429-432 (1992)
[Publications] Y.Akiyama et al.: "In vitro catalysis of oxidative folding of disulfide-bonded proteins by Escherichia coli dsdA (ppfA) gene product." J.Biol.Chem.267. 22440-22445 (1992)
[Publications] T.Mizobata te al.: "Effects of the chaperonin GroE on the refolding of tryptophanase from Escherichia coli.Refolding is enhanced in the presence of ADP." J.Biol.chem.267. 17773-17779 (1992)
[Publications] Y.Akiyama et al.: "Folding and assembly of bacterial alkaline phosphatase in vitro and in vivo." J.Biol.Chem.268.

1992 Fiscal Year Annual Research Report

大腸菌蛋白質分泌の分子機構

Principal Investigator

水島 昭二 東京大学, 応用微生物研究所, 教授 (50013313)

Research Products

[Publications] M.Kato et al.: "In Vitro translocation of secretory proteins possessing no charges at the mature domain takes place efficiently in a protonmotive force-dependent manner." J.Biol.Chem.267. 413-418 (1992)

[Publications] L.Bruncage et al.: "SecY,SecE and band 1 form the membrane-embedded domain of E.coli preprotrin translocase." J.Biol Chem.267. 4166-4170 (1992)

[Publications] C.Hikita et al.: "Effects of total-hydrophobicity and length of the hydrophobic domain of a signal peptied on in vitro translocation effciency." J.Biol Chem.267. 4882-4888 (1992)

[Publications] K.Nishiyama et al.: "The C-terminal region of SecE interacts with SecY and id functional in the reconstitution of protein translocation activity in Escherichia coli." J.Biol Chem.267. 7170-7176 (1992)

[Publications] S.Matsuyama et al.: "Overproduction,purification and characterization of SecD and SecF,integral membrane components of the protein translocation machinery of Escherichia coli." Biochim.Biophys.Acta. 1122. 77-84 (1992)

[Publications] C.Hikita et al.: "The requirement of a positive charge at the amino terminus can be compensated for by a longer central hydrophobic stretch in the functioning of signala peptides." J.Biol.Chem.267. 12375-12379 (1992)

[Publications] H.Takamatsu et al.: "In vivo and in vitro characterization of the SecA gene product of bacillus subtilis." J.Bacteriol.174. 4308-4316 (1992)

[Publications] S,Matsuyama et al.: "Large-scale Production of membrane proteins fused to a truncated SecA in Escherichia coli." Biosci.Biotech.Biochem.56. 1512-1514 (1992)

[Publications] S.Kamitani te al.: "Identification and characterization of an Escherichia coli gene required for the formation of correctly folded alkaline phosphatase,a periplasmic enzyme." BMBO J.11. 57-62 (1992)

[Publications] C.Ueguchi et al.: "Multicopy suppression:an approach to understanding intracellular functioning of the protein export system." J.Bacteriol.174. 1454-1461 (1992)

[Publications] T.Shimoike et al.: "SecY variants that interfere with Escherichia coll protein export in the presence of normal secY." Mol.Microbiol.6. 1205-1210 (1992)

[Publications] K,Ito et al.: "SecY and integral membrane components of the Escherichia colf protein translocation ststem." Mol.Microbiol.6. 2423-2428 (1992)

[Publications] T.Taura et al.: "Insertional disruption of the nusB gene leads to cold-sensitive growth and suppression of the secY24 mutation." Mol.Gen.Genet.234. 429-432 (1992)

[Publications] Y.Akiyama et al.: "In vitro catalysis of oxidative folding of disulfide-bonded proteins by Escherichia coli dsdA (ppfA) gene product." J.Biol.Chem.267. 22440-22445 (1992)

[Publications] T.Mizobata te al.: "Effects of the chaperonin GroE on the refolding of tryptophanase from Escherichia coli.Refolding is enhanced in the presence of ADP." J.Biol.chem.267. 17773-17779 (1992)

[Publications] Y.Akiyama et al.: "Folding and assembly of bacterial alkaline phosphatase in vitro and in vivo." J.Biol.Chem.268.

水島昭二東京大学, 応用微生物研究所, 教授 (50013313)