2001 Fiscal Year Annual Research Report

構造情報に基づく抗原抗体反応の精密解析と特異性・親和性を決定する因子の抽出

Research Project

Project/Area Number	12780486
Research Institution	Tohoku University
Principal Investigator	津本浩平東北大学, 大学院・工学研究科, 講師 (90271866)
Keywords	抗原抗体相互作用 / 蛋白性抗原 / X線結晶構造解析 / 熱力学的解析 / 分子認識 / 変異体 / 非共有結合 / 蛋白質間相互作用
Research Abstract	蛋白質間相互作用をより深く理解するために本研究では、モデルとなる蛋白性抗原と抗体可変領域の相互作用を取り上げ、(1)疎水的相互作用、(2)静電的相互作用(塩橋・水素結合)、(3)局所的構造変化に関与するアミノ酸残基、そして(4)界面に残存する溶媒分子に着目し、変異導入を施した抗原あるいは抗体を用いた相互作用について、構造解析・熱力学的解析を中心とした精密解析を行い、抗原抗体相互作用における特異性・親和性がどのような因子によって創出されているかを明らかにすることを目的としている。今年度の実績は以下の通りである。 (1)変異抗体と野生型抗原複合体の結晶構造解析:(1)水素結合を欠損させた軽鎖変異体(L-Asn31Ala, L-Asn31Asp, L-Asn32Ala, L-Asn32Asp,L-Asn92Ala, L-Asn92Asp)、(2)水素結合を欠損した重鎖変異体(H-Ser52Ala、H-Ser54Ala、H-Ser56Ala)、(3)抗原との誘導適合に重要な役割を果たしている重鎖Tyr53の側鎖を欠損させた変異体(H-Tyr53Ala)と野生型抗原との複合体について、熱力学的解析を詳細に行ったほか、構造解析に相応しい結晶を得、これらの結晶構造を精密化し、得られた構造情報を基に、熱力学的パラメーターの変化との関連を議論することができた。(2)三者複合体間の界面における水分子の役割の考察:変異抗体(L-Tyr50Phe、L-Ser91Ala、L-Tyr93Ala)との野生型抗原との相互作用について、熱力学的に解析し、複合体について、構造解析にふさわしい結晶を作製、結晶構造を精密化し、得られた構造情報に基づいて、熱力学的パラメーターとの変化を議論できた。(3)重鎖軽鎖間相互作用の役割:ファージ提示法により機能変換を施した変異抗体について、変異抗原ならびに野生型抗原との複合体の結晶構造を明らかにし、重鎖軽鎖間相互作用の抗原特異性への寄与について深く議論できた。

Research Products

(23 results)

All Other

All Publications (23 results)

[Publications] Yoda et al.: "Folding-unfolding of goat aipha-lactalbumin studied by stopped-flow CD and molecular dynamic simulations"Proteins ; Struct. Funct. Genet.. 42. 49-65 (2001)
[Publications] Arai et al.: "Demonstration of a homogeneous noncompetitive immunoassay based on bioluminescence resonance energy transfer"Analytical Biochemistry. 289. 77-81 (2001)
[Publications] Nakamura et al.: "Panning of phage VH library using nitrocellulose membranes-application to selection of human VH library"The Journal of Biochemistry. 129. 209-212 (2001)
[Publications] Kasai et al.: "Fv fragment of an antibody specific for hepatitis C NS3 protease domain : The sequence analysis, bacterial expression and functional characterization"Biochem. Biophys. Res. Commun.. 281. 416-424 (2001)
[Publications] Ueno et al.: "T-cell Immunotherapy for Human MK-1-Expressing Tumors Using a Fusion Protein of the Superantigen SEA and Anti-MK-1 scFv Antibody"Anticancer research. 22(印刷中). (2002)
[Publications] 津本: "不溶性顆粒からの蛋白質の巻き戻し"蛋白質核酸酵素. 46. 1238-1246 (2001)
[Publications] 津本, 熊谷: "ファージディスプレーを用いた新規バイオ分子の選択と応用"BIOINDUSTRY. 18. 26-33 (2001)
[Publications] 藤井, 津本: "抗体系-コンビナトリアルバイオエンジニアリング"現代化学. 369. 40-45 (2001)
[Publications] Shiroishi et al.: "Structural evidence for entropic contribution of salt bridge formation to a protein antigen-antibody interaction : The case of HyHEL-10 Fv- hen lysozyme complex"The Journal of Biological Chemistry. 276. 23042-23050 (2001)
[Publications] Kawahara et al.: "Replacing factor dependency with that for lysozyme : Affordable culture of IL-6 dependent hybridoma by transfecting artificial cell surface receptor"Biotechnology and Bioengineering. 76. 416-423 (2001)
[Publications] Nakamura et al.: "The effect of an agglutogen on virus infection : biotinylated filamentous phages and avidin as a model"FEBS Letters. (印刷中). (2002)
[Publications] Kawahara et al.: "Selection of highly productive mammalian cells based on an inducible growth advantage using an antibody/receptor chimera"J. Biosci. Biotechnol.. (印刷中). (2002)
[Publications] Kawahara et al.: "A growth signal with an artificially induced erythropoietin receptor-gp130 cytoplasmic domain heterodimer"The Journal of Biochemistry. 130. 305-312 (2001)
[Publications] Horii et al.: "Contribution of Thr29 to the Thermodynamic Stability of Goat α-Lactalbumin As Determined by Experimental and Theoretical Approaches"PROTEINS : Struct. Funct. Genet.. 45. 16-29 (2001)
[Publications] Nakamura et al.: "A visualization method of filamentous phage infection and phage-derived proteins in Eseherichia coli using biotinylated phages"Biochem. Biophys. Res. Commun.. 289. 252-256 (2001)
[Publications] Kodama et al.: "Mutated SEA D227A-conjugated antibodies greatly enhance antitumor activity against MUC1-expressing bile duct carcinoma(BDC)"Cancer Immunology Immunotherapy. 50. 539-548 (2001)
[Publications] Nakamura et al.: "Phage library panning against cytosolic fraction of cells using quantitative dot blotting : Application of selected VH to histochemistry"Journal of Immunological Methods. 261. 65-72 (2002)
[Publications] Takemura et al.: "A mutated superantigen SEA D227A fusion diabody specific to MUC1 and CD3 for targeted cancer immunotherapy for bile duct carcinoma"Cancer Immunology Immunotherapy. (印刷中). (2002)
[Publications] Nakamura et al.: "Detection of biotinylated proteins in polyacrylamide gels using an avidin-fluorescein conjugate"Analytical Biochemistry. (印刷中). (2002)
[Publications] Kumagai, Tsumoto: "Encyclopedia of Life Sciences"Blackwell(印刷中). (2002)
[Publications] 津本, 熊谷: "コンビナトリアルバイオエンジニアリング"化学同人(印刷中). (2002)
[Publications] 津本, 熊谷: "新タンパク質科学実験法第3巻構造機能解析"東京化学同人(印刷中). (2002)
[Publications] 津本, 熊谷: "生物物理学ハンドブック"朝倉書店(印刷中). (2002)

2001 Fiscal Year Annual Research Report

構造情報に基づく抗原抗体反応の精密解析と特異性・親和性を決定する因子の抽出

Principal Investigator

津本 浩平 東北大学, 大学院・工学研究科, 講師 (90271866)

Research Products

[Publications] Yoda et al.: "Folding-unfolding of goat aipha-lactalbumin studied by stopped-flow CD and molecular dynamic simulations"Proteins ; Struct. Funct. Genet.. 42. 49-65 (2001)

[Publications] Arai et al.: "Demonstration of a homogeneous noncompetitive immunoassay based on bioluminescence resonance energy transfer"Analytical Biochemistry. 289. 77-81 (2001)

[Publications] Nakamura et al.: "Panning of phage VH library using nitrocellulose membranes-application to selection of human VH library"The Journal of Biochemistry. 129. 209-212 (2001)

[Publications] Kasai et al.: "Fv fragment of an antibody specific for hepatitis C NS3 protease domain : The sequence analysis, bacterial expression and functional characterization"Biochem. Biophys. Res. Commun.. 281. 416-424 (2001)

[Publications] Ueno et al.: "T-cell Immunotherapy for Human MK-1-Expressing Tumors Using a Fusion Protein of the Superantigen SEA and Anti-MK-1 scFv Antibody"Anticancer research. 22(印刷中). (2002)

[Publications] 津本: "不溶性顆粒からの蛋白質の巻き戻し"蛋白質核酸酵素. 46. 1238-1246 (2001)

[Publications] 津本, 熊谷: "ファージディスプレーを用いた新規バイオ分子の選択と応用"BIOINDUSTRY. 18. 26-33 (2001)

[Publications] 藤井, 津本: "抗体系-コンビナトリアルバイオエンジニアリング"現代化学. 369. 40-45 (2001)

[Publications] Shiroishi et al.: "Structural evidence for entropic contribution of salt bridge formation to a protein antigen-antibody interaction : The case of HyHEL-10 Fv- hen lysozyme complex"The Journal of Biological Chemistry. 276. 23042-23050 (2001)

[Publications] Kawahara et al.: "Replacing factor dependency with that for lysozyme : Affordable culture of IL-6 dependent hybridoma by transfecting artificial cell surface receptor"Biotechnology and Bioengineering. 76. 416-423 (2001)

[Publications] Nakamura et al.: "The effect of an agglutogen on virus infection : biotinylated filamentous phages and avidin as a model"FEBS Letters. (印刷中). (2002)

[Publications] Kawahara et al.: "Selection of highly productive mammalian cells based on an inducible growth advantage using an antibody/receptor chimera"J. Biosci. Biotechnol.. (印刷中). (2002)

[Publications] Kawahara et al.: "A growth signal with an artificially induced erythropoietin receptor-gp130 cytoplasmic domain heterodimer"The Journal of Biochemistry. 130. 305-312 (2001)

[Publications] Horii et al.: "Contribution of Thr29 to the Thermodynamic Stability of Goat α-Lactalbumin As Determined by Experimental and Theoretical Approaches"PROTEINS : Struct. Funct. Genet.. 45. 16-29 (2001)

[Publications] Nakamura et al.: "A visualization method of filamentous phage infection and phage-derived proteins in Eseherichia coli using biotinylated phages"Biochem. Biophys. Res. Commun.. 289. 252-256 (2001)

[Publications] Kodama et al.: "Mutated SEA D227A-conjugated antibodies greatly enhance antitumor activity against MUC1-expressing bile duct carcinoma(BDC)"Cancer Immunology Immunotherapy. 50. 539-548 (2001)

[Publications] Nakamura et al.: "Phage library panning against cytosolic fraction of cells using quantitative dot blotting : Application of selected VH to histochemistry"Journal of Immunological Methods. 261. 65-72 (2002)

[Publications] Takemura et al.: "A mutated superantigen SEA D227A fusion diabody specific to MUC1 and CD3 for targeted cancer immunotherapy for bile duct carcinoma"Cancer Immunology Immunotherapy. (印刷中). (2002)

[Publications] Nakamura et al.: "Detection of biotinylated proteins in polyacrylamide gels using an avidin-fluorescein conjugate"Analytical Biochemistry. (印刷中). (2002)

[Publications] Kumagai, Tsumoto: "Encyclopedia of Life Sciences"Blackwell(印刷中). (2002)

[Publications] 津本, 熊谷: "コンビナトリアルバイオエンジニアリング"化学同人(印刷中). (2002)

[Publications] 津本, 熊谷: "新タンパク質科学実験法 第3巻構造機能解析"東京化学同人(印刷中). (2002)

[Publications] 津本, 熊谷: "生物物理学ハンドブック"朝倉書店(印刷中). (2002)

津本浩平東北大学, 大学院・工学研究科, 講師 (90271866)

[Publications] 津本, 熊谷: "新タンパク質科学実験法第3巻構造機能解析"東京化学同人(印刷中). (2002)