| Project/Area Number |
12470506
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
医薬分子機能学
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
KOIKE Tohru Hiroshima University, Graduate School of Biomedical Sciences, Professor, 大学院・医歯薬学総合研究科, 教授 (90186586)
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| Co-Investigator(Kenkyū-buntansha) |
OHTANI Kazuhiro Hiroshima University, Graduate School of Biomedical Sciences, Associate Professor, 大学院・医歯薬学総合研究科, 助教授 (20203820)
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| Project Period (FY) |
2000 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥8,000,000 (Direct Cost: ¥8,000,000)
Fiscal Year 2002: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2001: ¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2000: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Keywords | Intelligent molecule / Protease / Artificila Enzyme / Macrocyclic Polyamine / Phosphatase / 亜鉛酵素 / フォスフォプロテオーム |
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| Research Abstract |
Macrocyclic poly amines are a new class of multifunctional molecules (e.g., complexing agents for cations, anions and neutral molecules). Recently, a new insight into zinc enzyme chemistry has been achieved by studies of mononuclear and multinuclear zinc(II) complexes with macrocyclic polyamines. These complexes serve as both the structural and functional models for the active centers of zinc(II)-containing phosphatases (e.g., alkaline phosphatase, and protein serine/threonine phosphatase-1) and proteases (e.g., carboxypepetidase). Those study have given answers to some basic questions that surround the intrinsic properties of metal ions in those enzymes. On the bases of the finding from the zinc(II)-enzyme models, we synthesized novel artificial proteases and phosphatases (FW < 1000) and then characterized the compounds. In the course of an attempt to synthesize a new multinuclear zinc(II) complex for a phosphatase model, we incidentially isolated a new hexaaza cryptand dizinc(II) complex. The dinuclear zinc(II) complex (Phos-tag^<TM>) forms an extremely stable 1:1 phosphate complex in aqueous solution at physiological pH. We found that Phos-tag^<TM> recognizes phosphate monoester dianion (dissociation constat K_d = ca. 10^<-8> M) over 10^4 times selectively against carboxylate anions, which should facilitate further development of novel analytical methods for phospho-proteomics. As the first practical examples; we demonstrate the MADI-TOF mass spectrometry of phosphoryl-peptides by using Phos-tag^<TM> and a neutral matrix (e.g., THAP) at physiological pH. The single zinc-isotope Phos-tag^<TM> molecules (^<64>Zn and ^<68>Zn) enable the improvement of sensitivity and the analysis of phosphorylated residue.
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