Development of a method for membrane protein synthesis based on ligation chemistry

• Project/Area Number: 15083204
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: Osaka University
• Principal Investigator: AIMOTO Saburo Osaka University, 蛋白質研究所, 教授 (80029967)
• Co-Investigator(Kenkyū-buntansha): KAWAKAMI Toru 大阪大学, 蛋白質研究所, 准教授 (70273711) ; SATO Takeshi 大阪大学, 蛋白質研究所, 助教 (90403013)
• Project Period (FY): 2003 – 2008
• Project Status: Completed (Fiscal Year 2008)
• Budget Amount: ¥87,000,000 (Direct Cost: ¥87,000,000); Fiscal Year 2008: ¥16,000,000 (Direct Cost: ¥16,000,000); Fiscal Year 2007: ¥16,000,000 (Direct Cost: ¥16,000,000); Fiscal Year 2006: ¥15,500,000 (Direct Cost: ¥15,500,000); Fiscal Year 2005: ¥15,500,000 (Direct Cost: ¥15,500,000); Fiscal Year 2004: ¥16,000,000 (Direct Cost: ¥16,000,000); Fiscal Year 2003: ¥8,000,000 (Direct Cost: ¥8,000,000)
• Keywords: 膜蛋白質 / 化学合成 / ライゲーション法 / 補助基 / ペプチドチオエステル / 溶解性 / 合成法 / 膜近傍部位 / EGF受容体 / 電位依存性プロトンチャネル / EGF 受容体 / v-ATPase / Fmoc法 / 光反応性補助基 / 4回膜貫通型受容体 / 選択的縮合法 / Argタグ補助基

Research Abstract

ライゲーションケミストリーの開発を中心課題とし、膜蛋白質の合成法に関する総合的研究を行った。また、本研究の過程で得られた知見を元に、膜貫通ドメインを有するペプチドや蛋白質の合成を行い、FGFR3、ErbB2/Neuなどの受容体型チロシンキナーゼや、アミロイド前駆体蛋白質の膜貫通部位の構造解析を行った。また、四回膜貫通型蛋白質の化学合成に世界で始めて成功した。

Research Products

• [Journal Article] The use of a cysteinyl prolyl ester (CPE) autoactivating unit in peptide ligation reactions (2009)
  • Author(s): T. Kawakami, S. Aimoto
  • Journal Title: Tetrahedron 65 Pages: 3871-3877
  • Peer Reviewed
• [Journal Article] Peptide purification by affinity chromatography based on α-ketoacyl group chemistry (2009)
  • Author(s): T. Hara, A. Tainosho, K. Nakamura, T. Sato, T. Kawakami, S. Aimoto
  • Journal Title: J. Peptide Sci. 15 Pages: 369-376
  • Peer Reviewed
• [Journal Article] Structure of the transmembrane dimer of the amyloid precursor protein: Implications for proteolysis by g-secretase (2009)
  • Author(s): T. Sato, T.-C.Tang, G. Reubins, J.Z. Fei, T. Fujimoto, P. Kienlen-Campard, S. N. Constantinescu, J.-N. Octave, S. Aimoto, S. O. Smith
  • Journal Title: Proc. Natl Acad. Sci. USA 106 Pages: 1421-1426
  • Peer Reviewed
• [Journal Article] Evaluation of Lipid-Binding Properties of the N-Terminal Helical Segments in Human Apolipoprotein A-I Using Fragment Peptides (2009)
  • Author(s): M. Tanaka
  • Journal Title: J. Peptide Sci. 15 Pages: 36-42
  • Peer Reviewed
• [Journal Article] A Helix-to-Coil Transition at the ε-Cut Site in the Transmembrane Dimer of the Amyloid Precursor Protein is Reauired for Proteolvsis (2009)
  • Author(s): T. Sato
  • Journal Title: Proc.Natl.Acad.Sci.USA 106 Pages: 1421-1426
  • Peer Reviewed
• [Journal Article] Peptide purification by affinity chromatography based on α-ketoacyl group chemistry (2009)
  • Author(s): T. Hara
  • Journal Title: J. Pept. Sci. (In press)
  • Peer Reviewed
• [Journal Article] ^1H MAS RFDR NMR of biological membranes (2009)
  • Author(s): D. Aucoin
  • Journal Title: J. Magn. Reson (In press)
  • Peer Reviewed
• [Journal Article] 2-Nitrobenzylcarbamate-type Photocleavable Linker Having Fmoc-aminoalkyloxy Group : Preparation and Application to Peptide Purification (2009)
  • Author(s): T. Hara
  • Journal Title: Peptide Science 2008 (In press)
  • Peer Reviewed
• [Journal Article] Synthesis of Voltage-Dependent Proton Channel(VSOP)(121-222) (2009)
  • Author(s): K. Nakamura
  • Journal Title: Peptide Science 2008 (In press)
  • Peer Reviewed
• [Journal Article] The Use of a Cysteinyl Prolyl Ester(CPE)Autoactivating Unit in Peptide Ligati on Reactions (2009)
  • Author(s): T. Kawakami
  • Journal Title: Tetrahedron (In press)
  • Peer Reviewed
• [Journal Article] Amyloidogenic processing but not aicd production requires a precisely oriented APP dimer assembled by transmembrane GXXXG motifs (2008)
  • Author(s): P. Kienlen-Campard
  • Journal Title: J. Biol. Chem. 283 Pages: 7733-7744
  • Peer Reviewed
• [Journal Article] Roles of integral protein in membrane permeabilization by amphidinols (2008)
  • Author(s): N. Morsy
  • Journal Title: Biochim Biophys Acta 1778 Pages: 1453-1459
  • Peer Reviewed
• [Journal Article] Design and Synthesis of Ladder-shaped Tetracyclic, Heptacyclic, and Decacyclic Ethers and Evaluation of the Interaction with Transmembrane Proteins (2008)
  • Author(s): K. Torikai
  • Journal Title: J. Am. Chem. Soc. 130 Pages: 10217-10226
  • Peer Reviewed
• [Journal Article] Interaction of ladder-shaped polyethers with transmembrane a-helix of glycophorin A as evidenced by surface plasmon resonance and saturation transfer difference NMR (2008)
  • Author(s): S. Ujihara
  • Journal Title: Bioorg. Med. Chem. Lett. 18 Pages: 6115-8
  • Peer Reviewed
• [Journal Article] Amyloidogenic processing but not aicd production requires a precisely oriented APP dimer assembled by transmembrane GXXXG motifs. (2008)
  • Author(s): P., Kienlen-Campard
  • Journal Title: J. Biol. Chem. Epub(印刷中)
  • Peer Reviewed
• [Journal Article] Roles of integral protein in membrane permeabilization by amphidinols. (2008)
  • Author(s): N., Morsy
  • Journal Title: Biochim. Biophys. Acta Epub(印刷中)
  • Peer Reviewed
• [Journal Article] Peptide Ligation Using the Cysteinyl Prolyl Ester (CPE) Activating Unit at the Carboxy Terminus (2007)
  • Author(s): T. Kawakami, S. Aimoto
  • Journal Title: Chem. Lett. 36 Pages: 76-77
  • Peer Reviewed
• [Journal Article] Sequential peptide ligation by using a controlled cysteinyl prolyl ester (CPE) autoactivating unit. (2007)
  • Author(s): T., Kawakami
  • Journal Title: Tetrahedron Lett. 48 Pages: 1903-1905
  • Peer Reviewed
• [Journal Article] Peptide Ligation Using the Cysteinyl Prolyl Ester (CPE) Activating Unit at the Carboxy Terminus (2007)
  • Author(s): T.Kawakami
  • Journal Title: Chem. Lett. 36 Pages: 76-77
• [Journal Article] Peptide Thioester Synthesis via an Auxiliary-Mediated N-S Acyl Shift Reaction in Solution (2007)
  • Author(s): K.Nakamura
  • Journal Title: Int. J. Peptide Res. Therapeutics (E-pub)
• [Journal Article] Studies on substrate specificity at PR/p3 cleavage site of HTLV-1 protease (2007)
  • Author(s): J.K.Bang
  • Journal Title: Int. J. Peptide Res. Therapeutics (E-pub)
• [Journal Article] Generation of an S-Peptide via an N-S Acyl Shift Reaction in TFA Solution (2006)
  • Author(s): K. Nakamura, M. Sumida, T. Kawakami, T. Vorherr, S. Aimoto
  • Journal Title: Bull Chem Soc. Jpn. 79 Pages: 1773-1780
  • NAID: 10019373528
  • Peer Reviewed
• [Journal Article] Inhibitors of amyloid toxicity based on beta-sheet packing of Aβ40 and Aβ42. (2006)
  • Author(s): T. Sato, P. Kienlen-Campard, M. Ahmed, W. Liu, H. Li, J. I. Elliott, S. Aimoto, S. N. Constantinescu, J. N. Octave, S. O. Smith
  • Journal Title: Biochemistry 45 Pages: 5503-16
  • Peer Reviewed
• [Journal Article] Evaluations of Substrate Specificity and Inhibition at PR/p3 Cleavage Site of HTLV-1 Protease (2006)
  • Author(s): H.Naka
  • Journal Title: Bioorg. Med. Chem. Lett. 16 Pages: 3761-3764
• [Journal Article] Inhibitors of arnyloid toxicity based on beta-sheet packing of Abeta40 and Abeta42 (2006)
  • Author(s): T.Sato
  • Journal Title: Biochemistry 45 Pages: 5503-5516
• [Journal Article] Generation of an S-Peptide via an N-S Acyl Shift Reaction in TFA Solution (2006)
  • Author(s): K.Nakamura
  • Journal Title: Bull Chem Soc. Jpn. 79 Pages: 1773-1780
• [Journal Article] Peptide Thioester Preparation Based on an N-S Acyl Shift Reaction Mediated by a Thiol Ligation Auxiliary. (2005)
  • Author(s): T. Kawakami,M. Sumida, K. Nakamura, T. Vorherr, S. Aimoto
  • Journal Title: Tetrahedron Lett. 46 Pages: 8805-8807
  • Peer Reviewed
• [Journal Article] Synthesis of the C-terminal Region of Opioid Receptor Like 1 in an SDS Micelle by the Native Chemical Ligation: Effect of Thiol Additive and SDS Concentration on Ligation Efficiency, (2005)
  • Author(s): T. Sato, Y. Saito, S. Aimoto
  • Journal Title: J. Peptide Sci. 11 Pages: 410-416
  • Peer Reviewed
• [Journal Article] Sequential Peptide Chemical Ligation by the Thioester Method and Extended Chemical Ligation. (2005)
  • Author(s): T. Kawakami, M. Tsuchiya, K. Nakamura, S. Aimoto
  • Journal Title: Tetrahedron Lett. 46 Pages: 5533-5536
  • Peer Reviewed
• [Journal Article] Synthesis of the C-terminal Region of Opioid Receptor Like 1 in an SDS Micelle by the Native CChemical Ligation Effect of Thiol Additive and SDS Concentration on Ligation Efficiency (2005)
  • Author(s): T.Sato
  • Journal Title: J.Peptide Sci. 11 Pages: 410-416
• [Journal Article] Sequential Peptide Chemical Ligation by the Thioester Method and Extended Chemical Ligation (2005)
  • Author(s): T.Kawakami
  • Journal Title: Tetrahedron Lett. 46 Pages: 5533-5536
• [Journal Article] Peptide Thioester Preparation Based on an N-S Acyl Shift Reaction Mediated by a Thiol Ligation Auxiliary (2005)
  • Author(s): T.Kawakami
  • Journal Title: Tetrahedron Lett. 46 Pages: 8805-8807
• [Journal Article] Structural Role of Glycine in Amyloid Fibrils Formed from Transmembrane alpha-Helices (2005)
  • Author(s): W.Liu
  • Journal Title: Biochemistry 44 Pages: 3591-3597
• [Journal Article] Ladder-shaped polyether compound, desulfated yessotoxin, interacts with membrane-integral α-helix peptides (2005)
  • Author(s): M.Mori
  • Journal Title: Bioorganic & Medicinal Chemistry 13 Pages: 5099-5103
• [Journal Article] Switching of turn conformation in an aspartate anion peptide fragment by NH... O(-) hydrogen bonds. (2005)
  • Author(s): A.Onoda
  • Journal Title: Biopolymers, Epub. Jan Pages: 4-4
• [Journal Article] Synthesis of an olefin-containing cyclic peptide using the solid-phase Honer-Emmons reaction. (2004)
  • Author(s): J.K.Bang
  • Journal Title: Tetrahedron Lett. 45 Pages: 99-102
• [Journal Article] Intact Glycation End Products Containing Carboxylmethyl-Lysine and Glyoxal Lysine Dimer Obtained from Synthetic Collagen Model Peptide (2004)
  • Author(s): H.Yamada
  • Journal Title: Bioorg.Med.Chem.Lett. 14 Pages: 5677-5680
• [Journal Article] Synthesis of the C-terminal Region of Opioid Receptor Like 1 in an SDS Micelle by the Native Chemical Ligation : Effect of Thiol Additive and SDS Concentration on Ligation Efficiency. (2004)
  • Author(s): T.Sato
  • Journal Title: J.Peptide Sci., Epub. Dec. Pages: 20-20
• [Journal Article] Solid-Phase Staudinger Ligation on a Novel Core-Shell Type Resin
  • Author(s): H.Kim
  • Journal Title: Organic Lett. (In press)
• [Presentation] ネイティブケミカルライゲーション法とチオエステル法の併用を可能にするチオール保護基としてのメチルチオ基の利用 (2008)
  • Author(s): 赤井優一
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-30
• [Presentation] 電位依存性プロトンチャネル(VSOP)(121-222)の合成 (2008)
  • Author(s): 中村健一郎
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-30
• [Presentation] Synthesis of Peptide Thioester by the Cysteinyl Prolyl Ester(CPE)Method (2008)
  • Author(s): Toru Kawakami
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-30
• [Presentation] ヒトSerum Amyloid AのN末端領域は脂質結合に重要な役割を果たす (2008)
  • Author(s): 大田慎也
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-30
• [Presentation] アミノオキシ基とイソチオシアネート基の選択的化学反応とそれに続くエドマン分解を利用したペプチドの精製 (2008)
  • Author(s): 田結荘明
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-29
• [Presentation] Fmoc-アミノプロピルオキシ基を導入した2-ニトロベンジルカルバメイト型リンカーの合成とペプチド精製への応用 (2008)
  • Author(s): 原利明
  • Organizer: 第45回ペプチド討論会
  • Place of Presentation: 東京
  • Year and Date: 2008-10-29
• [Presentation] Peptide thioester preparation and peptide ligation using cysteinyl prolyl ester (CPE) autoactivating unit (2008)
  • Author(s): Toru Kawakami
  • Organizer: 30th European Peptide Symposium
  • Place of Presentation: ヘルシンキ
  • Year and Date: 2008-09-01
• [Presentation] Peptide thioester preparation and peptide ligation using cysteinyl prolyl ester(CPE)autoactivating unit (2008)
  • Author(s): Toru Kawakami
  • Organizer: 3oth European Peptide Symposium
  • Place of Presentation: ヘルシンキ
  • Year and Date: 2008-09-01
• [Presentation] N-4, 5-ジメトキシ-2-メルカプトベンジル基が介するN-Sアシル基転位反応を鍵反応としたペプチドチオエステル合成法の開発 (2008)
  • Author(s): 中村健一郎
  • Organizer: 日本化学会第88春季年会
  • Place of Presentation: 東京都
  • Year and Date: 2008-03-28
• [Presentation] α-ケトアシル基の形成と切断を鍵反応とするペプチド精製法の開発 (2008)
  • Author(s): 原利明
  • Organizer: 日本化学会第88春季年会
  • Place of Presentation: 東京
  • Year and Date: 2008-03-27
• [Presentation] N-Sアシル基転位反応を利用したペプチドチオエステルの調製 (2007)
  • Author(s): 中村健一郎
  • Organizer: 第44回ペプチド討論会
  • Place of Presentation: 富山市
  • Year and Date: 2007-11-07
• [Presentation] トロンボポエチン受容体の細胞外膜近傍部位-膜貫通部位の化学合成 (2007)
  • Author(s): 上杉知也
  • Organizer: 第44回ペプチド討論会
  • Place of Presentation: 富山市
  • Year and Date: 2007-11-07
• [Presentation] N to S Acyl Shift Reaction in Peptied Ligation (2007)
  • Author(s): T., Kawakami
  • Organizer: 4th international Peptied Symposium
  • Place of Presentation: オーストラリア・ケアンズ
  • Year and Date: 2007-10-23
• [Presentation] PEPTIDE THIOESTER SYNTHESIS BY AN ON-RESINAUXILIARY-MEDIATED N-S ACYL SHIFT REACTION (2007)
  • Author(s): S., Aimoto
  • Organizer: Modern Solid Phase Peptied Synthesis & Its Applications
  • Place of Presentation: オーストラリア・ポートダグラス
  • Year and Date: 2007-10-19
• [Presentation] Peptied Ligation via the N to S Acyl Shift Reaction (2007)
  • Author(s): T., Kawakami
  • Organizer: Chemical Protein Synthesis Meeting
  • Place of Presentation: オーストラリア・ヘロン島
  • Year and Date: 2007-10-18
• [Presentation] Peptied Ligation via the In-situ Transformation of an Amide in to a Thioester at a Cysteine Residue (2007)
  • Author(s): T., Kawakami
  • Organizer: American Peptied Symposium
  • Place of Presentation: カナダ・モントリオール
  • Year and Date: 2007-06-27
• [Remarks] ホームページ
  • URL: http://www.protein.osaka-u.ac.jp/organic/index.html
• [Patent(Industrial Property Rights)] ペプチドエステル試薬,およびそのライゲーションまたチオエステル化合物の製造のための使用 (2006)
  • Inventor(s): 相本三郎、川上徹
  • Industrial Property Rights Holder: 相本三郎、川上徹、大阪大学、大阪産業振興機構
  • Filing Date: 2006
  • Overseas
• [Patent(Industrial Property Rights)] ペプチドエステルを用いたライゲーション法、およびチオエステル化合物の製造方法 (2005)
  • Inventor(s): 川上徹、相本三郎
  • Industrial Property Rights Holder: 川上徹、相本三郎、大阪大学、大阪産業振興機構
  • Filing Date: 2005
• [Patent(Industrial Property Rights)] ペプチドエステルを用いたライゲーション法、およびチオエステル化合物の製造方法 (2005)
  • Inventor(s): 相本 三郎, 川上 徹
  • Industrial Property Rights Holder: 発明者大阪大学大阪産業振興機構
  • Filing Date: 2005-10-14
• [Publications] T.Kawakami, S.Aimoto: "A Photoremorable ligation auxiliary for use in polypeptide synthesis"Tetrahedron Letters. 44・32. 6059-6061 (2003)
• [Publications] T.Sato, S.Aimoto: "Use of thiosulfonate for the protection of thiol groups in peptide ligation by the thioester method."Tetrahedron Letters. 44・44. 8085-8087 (2003)