1991 年度実績報告書

タンパク質工学の基盤確立のための多角的アプロ-チ

研究課題

研究課題/領域番号	02304062
研究機関	九州大学
研究代表者	井本泰治九州大学, 薬学部, 教授 (90038282)
研究分担者	崎山文夫大阪大学, 蛋白質研究所, 教授 (40029947) 大塚栄子北海道大学, 薬学部, 教授 (80028836) 太田隆久東京大学, 農学部, 教授 (30011844) 岩永貞昭九州大学, 理学部, 教授 (90029942) 芦田玉一名古屋大学, 工学部, 教授 (10029936)
キーワード	タンパク質工学 / アルドラ-ゼ / NADHーシトクロムb5 / NMR / 血液凝固因子 / アロステリック現象 / T4エンドヌクレア-ゼ / ATPase.
研究概要	今年度もタンパク質工学の基盤を確立する目的で相互に密な連絡を取りながら以下の研究を行った。グリ-ンハブの4種のPLA_2アイソザイムを分離しアミノ酸配列を決めた(大野)。血液凝固VII因子中のSer52の水酸基に(Xyl)_2Glcがグリコシド結合していることを明らかにした(岩永)。ヒトアルドラ-ゼの7種のキメラタンパク質の触媒活性を詳しく解析した(堀)。T4エンドヌクレア-ゼのGlu23は、触媒活性に関与する残基であることを示した(大塚)。アミノアシルtRNA合成酵素の新しい活性測定法を開発した(外村)。アミノ酸変異を用いてH^+ATPaseのγサブユニットが、H^+輸送の制御を行っいることを明らかにした(二井)。枯草菌プロテア-ゼ分泌系を利用して、大腸菌細胞内酵素を分泌させることに成功した(鶴)。APIが高い触媒活性を示すのに、His210が重要であることを示唆した(崎山)。NADHーシトクロムb_5の変異体の構造と機能について検討した(指吸)。酵母とショウジョウバエ由来のRNAポリメラ-ゼIIの大サブユニットに原核生物由来のリボヌクレア-ゼとアミノ酸配列の類似性を持つ部分を見いだした(郷)。NMR及びジスタンスジオメトリ-アルゴリズムにより制ガン剤であるネオカルチノスタチンの高次構造を決定し、生理活性を持つクロモフォアの存在状態を議論した(小林)。ボ-マンバ-ク型プロテア-ゼインヒビタ-の結晶解析から、2つのドメインを構造的に維持するH_2Oの存在を見いだした(芦田)。Lー乳酸脱水素酵素におけるアロステリック現象は、リガンド結合によるわずかな3次構造変化により4次構造変化が誘起され、基質の親和性が変化することを明らかにした(太田)。新規に開発した糖鎖を有するタンパク質修飾試薬の付加によりRNaseT_1,Trypsinの熱安定性が増大することを示した(山崎)。リゾチ-ムのTrp108位のアミノ酸置換を行いGlu35の異常なpKa及び安定性に及ぼす効果を議論した(井本)。

研究成果
(24件)

すべてその他

すべて文献書誌 (24件)

[文献書誌] Ueda,Yamada,Sakamoto,Abe,Kawano,Terada and Imoto: "Preparation and Properties of a Lysozyme Derivative in Which Two Domains Are CrossーLinked Intramolecularly between Trp62 and Asp101." J.Biochem.110. 719-725 (1991)
[文献書誌] Koga,Yamada,Nishimura,Kato and Imoto: "Multiple Proteolytic Action of Rat Liver Cathepsin B:Specificities and pHーDependences of the Endoーand Exopeptidase Activities." J.Biochem.110. 179-188 (1991)
[文献書誌] Ueda,Isakari,Aoki,Yasukochi,Masutomo,Kawano,Terada,Yamada and Imoto: " ^1HーNMR Study of the Intramolecular Interaction of a Substrate Analogue Attached to Aspartic Acidー101 in Lysozyme." J.Biochem.109. 690-698 (1991)
[文献書誌] Koga,Yamada,Nishimura,Kato and Imoto: "Comparative Study in Specificities between Rat Cathepsin L and Papain:Amino Acid Differences at SubstrateーBinding Site Are Involved in Their Specificities." J.Biochem.109. 976-982 (1991)
[文献書誌] Suehiro,Miyata,Takeya,Takamatsu and Iwanaga: "Blood Clotting Factor IX Nagoya 3: The Molecular Defect of Zymogen Activation Caused by an Arginineー145 to Histidine一Substitution." Thrombosis Res.60. 311-320 (1991)
[文献書誌] Muta,Toshiyuki,Misumi,Tokunaga,Nakamura,Yoshihiro,Ikehara and Iwanaga: "Limulus Clotting Factor(Factor C):A Novel Protease that has a Mosaic Structure with Complementーlike and Lectinーlike Domains." J.Biol.Chem.266. 6554-6561 (1991)
[文献書誌] Tamaoki,Kobayashi,Nishimura,Ohkubo,Kyogoku,Nakajima,Kumagaye,Kimura and Sakakibara: "Solution conformational endothelin determined by means of ^1HーNMR spectroscopy and distance geometry calculations." Prot.Engineering.4. 509-518 (1991)
[文献書誌] Shirai and Go: "RNaseーlike domain in DNAーdirected RNA polymerase II." Proc.Natl.Acad.Sci.U.S.A.88. 9056-9060 (1991)
[文献書誌] Takagi,Omura and Go: "Evolutionary Origin of Thyroglobulin by Duplication of Esterase Gene." FEBS LETTERS. 282. 17-22 (1991)
[文献書誌] Yamane,Kobuke,Tsutsui,Toida,Suzuki,Ashida,Kawata and Sakiyama: "Crystal structure of Streptomyces ertyraeus trypsin at 2.7A resolution" J.Biochem.110. 945-951 (1991)
[文献書誌] Volbeda,Lahm,Sakiyama and Suck: "Crystal structure of Penicillium citrinum P1 nulclease at 2.8A resolution." EMBO J.10. 1607-1618 (1991)
[文献書誌] Kobayashi,Yoshimoto and Tsuru: "Chemical modification of neutral protease from Bacillus subtilis var.amylosacchariticus:assignment of tyrosyl residues iodinated." Agric.Biol.Chem.55. 2751-2756 (1991)
[文献書誌] Morikawa,Kanatani,Yoshimoto and Tsuru: "Dyeーsensitized photoxidation of neutral protease from B.subtilis var.amylosacchariticus:assignment of histidine residue oxidized." Agric.Biol.Chem.55. 2099-2104 (1991)
[文献書誌] Yamashita,Natatani and Tonomura: "SubstrataーSelective Activation of HistidineーModified Porcine Pancreatic αーAmylase by Chloride Ion." J.Biochem.110. 605-607 (1991)
[文献書誌] Iwamoto,Omoto,Hanada,Tomioka,Itai,Maeda and Futai: "Mutation in Serー174 and the glycineーrich sequence(Glyー149,Glyー150 and Thrー156)in the β subunit of Escherichia coli H^+ATPase." J.Biol.Chem.266. 16350-16355 (1991)
[文献書誌] Inahara,Iwamoto,Moriyama,Shimomura,Maeda and Futai: "Two genes,atpC1 and atpC2 for the γ subunit of Arabidopsis thaliana chloroplast ATP synthtase." J.Biol.Chem.266. 7333-7338 (1991)
[文献書誌] Nishida,Takahashi,Takahashi,Hisatsugu and Hori: "Monoclonal antibodies recognizing aminoーterminal and carboxyterminal regions of human aldolase A:Probes to detect conformational changes of the enzyme" J.Biochem.109. 544-550 (1991)
[文献書誌] Yamasaki,Kashina,Ohba and Funatsu: "Spectroscopic Analysis of the Interaction of Ricin D with Urea." Agric Biol.Chem.54. 1987-1994 (1991)
[文献書誌] Shirabe,Yubisui,Nishio and Takeshita: "Role of Cysteine Residues in Human NADHーcytochrome b_5 Reductase Studied by Siteーdirected Mutagenesis." J.Biol.Chem.266. 7531-7536 (1991)
[文献書誌] Imoto,Ueda,Tamura,Isakari,Inoue,Yasukochi,Miki,Horiuchi,Kawano and Yamada: "Enzyme Reaction Requires a Certain Fluctuation of the Active Site" Science.
[文献書誌] Ueda,Yamada and Imoto: "The Effect of Artificail Intramolecular CrossーLinking on the Renatuation Rate of Reduced Lysozyme." J.Biol.Chem.
[文献書誌] Ueda.Yamada,Nakashima,Tamura and Imoto: "The Detection of the Interaction in the Early Stage of Reduced Lysozyme by Use of Its Peptide Fragments." J.Biol.Chem.
[文献書誌] 井本泰治(分担): "タンパク質核酸酵素臨時増刊(タンパク質工学の進展)" 共立出版株式会社, 8 (1992)
[文献書誌] 山崎信行(分担): "新実験化学講座1 タンパク質IV 構造活性相関日本生化学会編" 東京化学同人, 28 (1991)

1991 年度 実績報告書

タンパク質工学の基盤確立のための多角的アプロ-チ

研究代表者

井本 泰治 九州大学, 薬学部, 教授 (90038282)

研究成果

[文献書誌] Ueda,Yamada,Sakamoto,Abe,Kawano,Terada and Imoto: "Preparation and Properties of a Lysozyme Derivative in Which Two Domains Are CrossーLinked Intramolecularly between Trp62 and Asp101." J.Biochem.110. 719-725 (1991)

[文献書誌] Koga,Yamada,Nishimura,Kato and Imoto: "Multiple Proteolytic Action of Rat Liver Cathepsin B:Specificities and pHーDependences of the Endoーand Exopeptidase Activities." J.Biochem.110. 179-188 (1991)

[文献書誌] Ueda,Isakari,Aoki,Yasukochi,Masutomo,Kawano,Terada,Yamada and Imoto: " ^1HーNMR Study of the Intramolecular Interaction of a Substrate Analogue Attached to Aspartic Acidー101 in Lysozyme." J.Biochem.109. 690-698 (1991)

[文献書誌] Koga,Yamada,Nishimura,Kato and Imoto: "Comparative Study in Specificities between Rat Cathepsin L and Papain:Amino Acid Differences at SubstrateーBinding Site Are Involved in Their Specificities." J.Biochem.109. 976-982 (1991)

[文献書誌] Suehiro,Miyata,Takeya,Takamatsu and Iwanaga: "Blood Clotting Factor IX Nagoya 3: The Molecular Defect of Zymogen Activation Caused by an Arginineー145 to Histidine一Substitution." Thrombosis Res.60. 311-320 (1991)

[文献書誌] Muta,Toshiyuki,Misumi,Tokunaga,Nakamura,Yoshihiro,Ikehara and Iwanaga: "Limulus Clotting Factor(Factor C):A Novel Protease that has a Mosaic Structure with Complementーlike and Lectinーlike Domains." J.Biol.Chem.266. 6554-6561 (1991)

[文献書誌] Tamaoki,Kobayashi,Nishimura,Ohkubo,Kyogoku,Nakajima,Kumagaye,Kimura and Sakakibara: "Solution conformational endothelin determined by means of ^1HーNMR spectroscopy and distance geometry calculations." Prot.Engineering.4. 509-518 (1991)

[文献書誌] Shirai and Go: "RNaseーlike domain in DNAーdirected RNA polymerase II." Proc.Natl.Acad.Sci.U.S.A.88. 9056-9060 (1991)

[文献書誌] Takagi,Omura and Go: "Evolutionary Origin of Thyroglobulin by Duplication of Esterase Gene." FEBS LETTERS. 282. 17-22 (1991)

[文献書誌] Yamane,Kobuke,Tsutsui,Toida,Suzuki,Ashida,Kawata and Sakiyama: "Crystal structure of Streptomyces ertyraeus trypsin at 2.7A resolution" J.Biochem.110. 945-951 (1991)

[文献書誌] Volbeda,Lahm,Sakiyama and Suck: "Crystal structure of Penicillium citrinum P1 nulclease at 2.8A resolution." EMBO J.10. 1607-1618 (1991)

[文献書誌] Kobayashi,Yoshimoto and Tsuru: "Chemical modification of neutral protease from Bacillus subtilis var.amylosacchariticus:assignment of tyrosyl residues iodinated." Agric.Biol.Chem.55. 2751-2756 (1991)

[文献書誌] Morikawa,Kanatani,Yoshimoto and Tsuru: "Dyeーsensitized photoxidation of neutral protease from B.subtilis var.amylosacchariticus:assignment of histidine residue oxidized." Agric.Biol.Chem.55. 2099-2104 (1991)

[文献書誌] Yamashita,Natatani and Tonomura: "SubstrataーSelective Activation of HistidineーModified Porcine Pancreatic αーAmylase by Chloride Ion." J.Biochem.110. 605-607 (1991)

[文献書誌] Iwamoto,Omoto,Hanada,Tomioka,Itai,Maeda and Futai: "Mutation in Serー174 and the glycineーrich sequence(Glyー149,Glyー150 and Thrー156)in the β subunit of Escherichia coli H^+ATPase." J.Biol.Chem.266. 16350-16355 (1991)

[文献書誌] Inahara,Iwamoto,Moriyama,Shimomura,Maeda and Futai: "Two genes,atpC1 and atpC2 for the γ subunit of Arabidopsis thaliana chloroplast ATP synthtase." J.Biol.Chem.266. 7333-7338 (1991)

[文献書誌] Nishida,Takahashi,Takahashi,Hisatsugu and Hori: "Monoclonal antibodies recognizing aminoーterminal and carboxyterminal regions of human aldolase A:Probes to detect conformational changes of the enzyme" J.Biochem.109. 544-550 (1991)

[文献書誌] Yamasaki,Kashina,Ohba and Funatsu: "Spectroscopic Analysis of the Interaction of Ricin D with Urea." Agric Biol.Chem.54. 1987-1994 (1991)

[文献書誌] Shirabe,Yubisui,Nishio and Takeshita: "Role of Cysteine Residues in Human NADHーcytochrome b_5 Reductase Studied by Siteーdirected Mutagenesis." J.Biol.Chem.266. 7531-7536 (1991)

[文献書誌] Imoto,Ueda,Tamura,Isakari,Inoue,Yasukochi,Miki,Horiuchi,Kawano and Yamada: "Enzyme Reaction Requires a Certain Fluctuation of the Active Site" Science.

[文献書誌] Ueda,Yamada and Imoto: "The Effect of Artificail Intramolecular CrossーLinking on the Renatuation Rate of Reduced Lysozyme." J.Biol.Chem.

[文献書誌] Ueda.Yamada,Nakashima,Tamura and Imoto: "The Detection of the Interaction in the Early Stage of Reduced Lysozyme by Use of Its Peptide Fragments." J.Biol.Chem.

[文献書誌] 井本 泰治(分担): "タンパク質核酸酵素臨時増刊(タンパク質工学の進展)" 共立出版株式会社, 8 (1992)

[文献書誌] 山崎 信行(分担): "新実験化学講座1 タンパク質IV 構造活性相関 日本生化学会編" 東京化学同人, 28 (1991)

1991 年度実績報告書

井本泰治九州大学, 薬学部, 教授 (90038282)

[文献書誌] 井本泰治(分担): "タンパク質核酸酵素臨時増刊(タンパク質工学の進展)" 共立出版株式会社, 8 (1992)

[文献書誌] 山崎信行(分担): "新実験化学講座1 タンパク質IV 構造活性相関日本生化学会編" 東京化学同人, 28 (1991)