| Project/Area Number |
17205015
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemistry related to living body
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| Research Institution | Hokkaido University |
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Principal Investigator |
NISIMURA Shin-ichitro Hokkaido University, Graduate School of Advanced Life Science, Professor (00183898)
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| Co-Investigator(Kenkyū-buntansha) |
NAGAHORI Noriko Hokkaido University, Graduate School of Advanced Life Science, Assistant Professor (90372268)
HINOU Hiroshi Hckkaido University, Graduate School of Advanced Life Science, Assistant Professor (70333333)
篠原 康郎 北海道大学, 大学院・理学研究科, 特任助教授 (20374192)
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| Project Period (FY) |
2005 – 2007
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| Project Status |
Completed (Fiscal Year 2007)
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| Budget Amount *help |
¥48,880,000 (Direct Cost: ¥37,600,000、Indirect Cost: ¥11,280,000)
Fiscal Year 2007: ¥13,260,000 (Direct Cost: ¥10,200,000、Indirect Cost: ¥3,060,000)
Fiscal Year 2006: ¥15,210,000 (Direct Cost: ¥11,700,000、Indirect Cost: ¥3,510,000)
Fiscal Year 2005: ¥20,410,000 (Direct Cost: ¥15,700,000、Indirect Cost: ¥4,710,000)
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| Keywords | gold colloidal nanoparticle / carbohydrate / glycoblotting / suicide substrate / MALDI-TOF MS / glycomics / quantum dots / live cell imaging / carbohydrate binding protein / proteomics / glycosyltransferase / mass spectrometry / enzyme assay / glycosidase / gold collodial nanoparticle |
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| Research Abstract |
this research was performed to understand the functions of glycoconjugates at the molecular level, that is to say, to develop a new methodology for hunting and identifying the partner molecules with target carbohydrates by using the gold colloidal nanoparticle and the mass spectrometer.
<Preparation water -soluble and non-specific adsorption depression type gold colloidal nanoparticle displaying carbohydrates>
in this study, the modification of nanoparticles was performed using specific affinity with a metallic surface and thiol group. Strong interaction of carbohydrate-displaying nanoparticle with a partner molecule is expected, because several hundred of carbohydrate molecules are displayed onto a surface of nanoparticle (carbohydrate cluster effect). In addition, the system that could display any carbohydrates was built by using chemoselective reaction with aminooxy1 group and a reducing end of carbohydrate. A library of carbohydrate presentation nano particle was made using this syst
… More
em. In addition, the surfaces of the nanoparticles were modified with suicide substrate type carbohydrate derivatives that could anchor the target proteins with covalent bonds. The presentation of carbohydrates on the nanoparticles was evaluated by MALDI-TOF MS using that the laser cleaved metal-thiol bond. In order to reduce the non-specific adhesion of proteins, a thiol compound with zwitterionic head group was designed and synthesized. All carbohydrate-displaying metal nanoparticles prepared in this study showed high water solubility and stability.
<Establishment of the methods for hunting and identifying the partner molecules of carbohydrates>
The highly water-soluble nanoparticles could be directly subjected to a various biochemical assays and MALDI-TOF MS measurement after the capture of partner molecules. Incubate nanoparticle and sample solution containing partner molecules, and wash the particle with buffer, then the particle was just applied to SDS-PAGE. The protein identification was possible according to the ordinary method of proteomics. In this way, a high sensitivity by reduced sample loss and a promotion of efficiency by simplification of activity process were achieved, compared with traditional approaches such as the affinity chromatography which needed elution by a competing carbohydrate. Less
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