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- きみのしん おおばま
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1 PHF (paired helical filament) Tau2r3 Fab2r3 MBD 1 MBD (1) The microtubule-binding domain (MBD) of tau protein is the region responsible for the PHF formation. We succeed in the preparation of an antibody Tau2r3, which selectively recognizes the MBD. The X-ray crystal structure analysis of the complex crystal of its variable region, Fab2r3, with the VQIIK fragment peptide of MBD clarified the overall complex structure. ow, we are investigating the recognition mode of MBD by the antibody at the atomic level. (2) The MBD interaction of a series of inhibitors we discovered so far were investigated by 1H-MR and other physicochemical methods, and clarified the structural elements necessary for exhibiting the inhibitory activity, i.e., inhibitor has a planar aromatic ring including the polar atom ( or ) or polar group (H or ) on the ring. n the other hand, the PHF formation experiment of a series of tau mutants clarified the important of CH π interaction between Ile308 and Tyr310 residues in the third repeat of MBD.
2 (a) (b) 1 R 1 R 2 R 3 R R2 microtubule-binding domain (MBD) Q T A P V P M P D L K V K K I G T E L K H Q P G G G K V Q I I K K L D L V Q K C G K D I K H V P G G G V Q I V Y K P V D L K V T K C G L G I H H K P G G G Q V E V K E K L D F K D R V Q K I G L D I T H V P G G G
3 体構造の８割以上の立体構造を同定してい る 本年中に精密な構造解析を完成させ 本 目的であるタウ抗体によるタウ MBD の選択 特異的認識機構を原子レベルで解明し この 情報を基に新規タウ重合阻害剤の開発に取 り組んでいる チオニン シアニジンをはじめとするＰＨ Ｆ形成阻害分子とタウＭＢＤドメインと の相互作用解析と それに基づく阻害分子 の開発と医薬品への展開 この知見に基づき チオニンより強いＰＨ Ｆ形成阻害能を示す天然分子ジュグロンお よびシコニンの発見に成功し 医薬への応 用のための分子改良を試み 現在 鋭意研 究を進めている Ｒ３ドメイ ン内チロ シン残 基を選択 特異 的に修飾する化合物の開発 一連のタウ蛋白質の変換体を作成し 変換 体残基の PHF 形成との構造活性相関について 研究した その結果 タウ蛋白質のＰＨＦ形 成に不可欠な構造要素として MBD 中の リピ ート内の３０８番目のイソロイシンと３１０ 番目のチロシン残基との間で形成される C-H π 結合が PHF 形成に不可欠であることを 明らかにした 一方 これまでに明らかにしてきたＰＨＦ形 成反応を阻害する低分子化合物を用いて そ の C-H π 結合を選択特異的にブロックする か否かについて MR をはじめとする分光化学 的奉手法により C-H π 結合阻害と PHF 形 成抑制との間に相関性のあることを明らかに した これまで当研究室で見出したチオニン シアニジンをはじめとする一連のタウ蛋白 のPHF形成阻害分子のＭＢＤとの相互作用 様式について 1Ｈ ＮＭＲ測定による溶液 構造解析および分子動力学計算および分光 化学的手法により解析を行った その結果 一連の阻害分子が R2-リピート分子や MBDの分子会合を阻害するために必要な 構造特徴として 芳香性の平面構造を有し てい ると と もにR2- リピ ート やMBDと 水素結合や静電結合が形成できる極性原子 を芳香環中または環に直接結合しているこ とが不可欠であることを明らかにすると共 に これにより MBD分子のβ シート構 造の形成を阻害できることを明らかにした Full-length Tau 306 R microtubul e-binding domain (MBD) 4RMBD 3RMBD Cyanidin R2 (-)-Epicatechin (-)-Catechin + His-tag Methylene blue Chlorpromazine Thionin + + Cl H2 H
4 10 K. Minoura, K. kuyama, C. ishiura, F. Mi zushima, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Role of each peptide in aggregatio n of tau microtubule-binding domain, Peptide cience, 2007, 2008, K. kuyama, C. ishiura, F. Mizushima, K. Minoura, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Linkage-dependent contribution of repeat peptides to self-aggregation of three- or four-repeat microtubule-binding domains in tau protein, FEB Journal,, 275, 2008, M. Hattori, E. ugino, K. Minoura, Y. In, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Different inhibitory response of cyanidin and methylene blue for filament formation of tau microtubule-binding domain, Biochem. Biophys. Res. Commun., 374, 2008, C. ishiura, K. Takeuchi, K. aruto, K. Minoura, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Importance of tyrosine residue for filament formation of tau microtubule-binding domain, Peptide cience, 2008, 2009, E. ugino, M. Hattori, K. Minoura, Y. In, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Inhibitory behavior of cyanidine and methylene blue for filament formation of tau microtubule-binding domain, Peptide cience,, 2008, 2009, E. ugino, C. ishiura, K. Minoura, Y. In, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Three-/four repeat-dependent aggregation profile of tau microtubule-binding domain clarified by dynamic light scattering analysis, Biochem. Biophys. Res. Commun.,, 385, 2009, T. Ishida, tructural studies of specific intermolecular interactions and self-aggregation of biomolecules and their application to drug design, Chem.Pharm.Bull., 57, 2009, C.ishiura, K. Takeuchi, K. Minoura, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, Importance of Tyr310 residue in the third repeat of microtubule binding domain for filament formation of tau protein, J. Biochem.,, 147, 2010, K. aruto, K. Minoura, R. kuda, T. Taniguchi, Y. In, T. Ishida, K. Tomoo, Interplay between I308 and Y310 residues in the their repeat of mucrotubule-binding domain is essential for tau filament formation, FEB Lett.,, 584, 2010, K. Minoura, K. Takeuchi, C. ishiura, K. aruto, M. umida, T. Taniguchi, K. Tomoo, T. Ishida, tructural role of tyrosine residue in the their repeat of microtubule binding domaion for filament formation of tau protein, Peptide cience,, 2009, 2010, , tau,, 20 08,, Tau (MBD)Tyrosine,, 2008,, Tau Tyrosinw,, 2008,, Cyanidine(Cy)Methy lene Blue(MB)Tau,, 2008,, t au,, 2008,
5 , Tau,, 2010,, Tau (MBD) Tyrosine,, 2010,, Tau (MBD)CH,, 2010,, Full length t au CH,, 2010, 1,,, 2010, 176
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