| Project/Area Number |
23510265
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Living organism molecular science
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| Research Institution | Nigata University of Phermacy and Applied Life Sciences |
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Principal Investigator |
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| Co-Investigator(Kenkyū-buntansha) |
ASADA Shinichi 新潟薬科大学, 薬学部, 助教 (50424883)
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| Project Period (FY) |
2011-04-28 – 2015-03-31
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| Project Status |
Completed (Fiscal Year 2014)
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| Budget Amount *help |
¥5,460,000 (Direct Cost: ¥4,200,000、Indirect Cost: ¥1,260,000)
Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2012: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2011: ¥2,340,000 (Direct Cost: ¥1,800,000、Indirect Cost: ¥540,000)
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| Keywords | 硫酸化タンパク質 / モノクローナル抗体 / ファージディスプレイ / チロシン硫酸化酵素 / 基質配列モチーフ / ペプチド合成 / プロテオーム解析 / 基質認識機構 / 固相ペプチド合成 / チオエステル縮合法 / クローニング / 硫酸化モチーフ / 生体分子 / 硫酸化ペプチド / 固相ペプチド化学合成 / 硫酸基転移酵素 / 硫酸化酵素 |
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| Outline of Final Research Achievements |
We have aimed to obtain the monoclonal antibody against sulfated tyrosine as immuno-chemical tool applicable to the sulfo-proteomics studies; however specific antibody against sulfated proteins could not be raised by the conventional immunization protocol in mice due to the inherent instability of sulfate. Thus we changed the approach to the phage-display method and now fundamental studies are in progress.
We have established the expression system in E. coli for two enzymes (TPST-1 and -2) responsible for tyrosine sulfation in protein. Using obtained proteins having enzyme activity and synthetic peptides based on ADYAE sequence, we found out the XDY (X: various amino acids) motif as a minimal peptide substrate for sulfation. We have developed a preparation procedure for peptide thioester containing tyrosine sulfate residue, and using the Ag+-mediated thioester segment condensation, we obtained the long-chain peptides containing multiple tyrosine sulfate residues in high yields.
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