Analysis of O-sulfotyrosine-mediated bio-interactions using synthetic peptides
| Project/Area Number |
13672241
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemical pharmacy
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| Research Institution | Nigata University of Phermacy and Applied Life Sciences |
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Principal Investigator |
KITAGAWA Kouki Niigata University of Pharmacy and Applied Life Sciences, Faculty of Pharmaceutical Sciences, Professor, 薬学部, 教授 (60093853)
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| Co-Investigator(Kenkyū-buntansha) |
関川 由美 新潟薬科大学, 薬学部, 助手 (50329330)
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| Project Period (FY) |
2001 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2003: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2002: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2001: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | Protein Sulfation / Tyrosine Sulfate / Chemical Synthesis / Solid-Phase Peptide Synthesis / Cholecystokinin / Thioester Condensation / α-Conotoxin / Tyrosin Sulfate Cluster / ペプチド性貝毒 / ジスルフィド結合 / 質量分析 / 硫酸化チロシン含有ペプチド |
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| Research Abstract |
1 We carried out the chemical synthesis of large molecular forms of Tyr(SO_3H)-containing peptides based on the facile and efficient solid-phase method developed by us.
i) Human big gastrin-II and its C-terminal Gly-extended peptide were prepared by the convergent segment condensation approach on the resin.
ii) Human cholecystokinin (CCK)-58 was prepared by the silver-ion mediated thioester segment condensation approach. In this synthesis, the 2-chlorotrityl resin was extensively used as a solid support to prepare the sulfated segment and partially protected thioester segments. CCK-58 exhibited glucose-dependent insulinotropic activity at levels comparable to CCK-33.
2 Three α-conotoxins (PnIA, PnIB, and EpI) that contain the sulfated tyrosine residue were synthesized by the Fmoc-based solid-phase method. Both a simultaneous oxidation approach and a two-step selective oxidation approach were employed to form the two disulfide linkages. In a simultaneous approach for PnIA and PnIB, additio
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n of DMSO in the reaction medium was critical to reduce the production of disulfide bond isomers. Desulfation was kept minimum throughout synthesis. α-Conotoxin EpI was synthesized using a novel solid-phase approach, in which α-carboxyl function of Asp was utilized as an anchoring group with a solid support. Three sulfated α-conotoxins were subjected to bioassay (Inhibitory effects of catecholeamine secretion from bovine adrenal chromaffin cells) and were found to be equipotent with their non-sulfate counterparts. This implies that the sulfate groups on these α-conotoxins are not determinants for their biological activities. Also we found that the disulfide bond isomers of these α-conotoxins had a weak but an apparent inhibitory effects of catecholeamine secretion.
3 Various sized rat CCK-peptides (CCK-33, CCK-22, and CCK-12) were prepared by our solid-phase method and they were used as markers on the HPLC separations of the proteolytic products from proCCK. A model for the progression of pro-CCK processing in AtT 20-cells was proposed.
4 Using the synthetic sulfated peptides, factor XI (FXI) was found to bind with the platelet glycoprotein Ibα not via an anionic duster in which three sulfated tyrosine residues were involved, but via the leucine-rich repeat sequences within the N-terminal domain of Gplbα. Less
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Report
(4 results)
Research Products
(23 results)
