| Project/Area Number |
12470487
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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 |
Physical pharmacy
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| Research Institution | Yokohama National University |
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Principal Investigator |
KATAHIRA Masato Yokohama National University, Faculty of Environment and Information Sciences, Associate Professor, 大学院・環境情報研究院, 助教授 (70211844)
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| Co-Investigator(Kenkyū-buntansha) |
UESUGI Seiichi Yokohama National University, Faculty of Environment and Information Sciences, Professor, 大学院・環境情報研究院, 教授 (70028851)
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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 |
¥8,400,000 (Direct Cost: ¥8,400,000)
Fiscal Year 2002: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 2001: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2000: ¥3,400,000 (Direct Cost: ¥3,400,000)
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| Keywords | HIV / aptamer / Tat protein / NMR / three-dimensional structure / RNA-binding protein / RNA / drug design / トリプレットリピート / リボザイム / ダナミクス / テロメア / 4重鎖 / 安定同位体標識 / 水素結合 / Tat蛋白質 / 蛋白質-RNA相互作用 |
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| Research Abstract |
An RNA aptamer containing two binding sites exhibits extremely high affinity to the HIV Tat protein. We have now determined the structure of the aptamer complexed with argininamide, the simplest analogue of the Tat protein, by NMR. We found that two argininamide molecules bind per aptamer. Two adjacent U : A : U base triples were formed, which widens the major groove to make space for the two argininamide molecules. The argininamide molecules bound to the G bases through hydrogen bonds. The binding is stabilized through stacking interactions.
The structure of the aptamer complexed with a Tat-derived argmine-rich peptide was also characterized. The Tat-derived peptide bound to the aptamer in a 1: 1 molar ratio. In spite of the difference in stoichiometry, similarity was noted between the argininamide- and peptide-bound forms of the aptamer in chemical shift perturbations of the aptamer upon complex formation and intermolecular contacts. These results suggest that two different arginine residues of the peptide interact with the two binding sites of the aptamer in the same way as two argininamide molecules do. Simultaneous interactions of the aptamer with two arginine residues of Tat could explain its high affinity compared to the authentic TAR RNA. The formation of the two adjacent base triples makes the simultaneous interactions possible by creating space for the accommodation of two arginine residues and linking residues of Tat, and also contributes to the stabilization of the binding through stacking interactions.
The elucidated structure of the aptamer gives clues to rationally design a new aptamer that exhibits even higher affinity to Tat. The way to design a new aptamer with a less side effect is also provided by the elucidated structure.
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