| Project/Area Number |
14380319
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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 |
Biophysics
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| Research Institution | Osaka University |
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Principal Investigator |
NAKAGAWA Atsushi Osaka University, Institute for Protein Research, Professor, 蛋白質研究所, 教授 (20188890)
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| Co-Investigator(Kenkyū-buntansha) |
YAMASHITA Eiki Osaka University, Institute for Protein Research, Instructor, 蛋白質研究所, 助手 (00294132)
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| Project Period (FY) |
2002 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥14,700,000 (Direct Cost: ¥14,700,000)
Fiscal Year 2005: ¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2004: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2003: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2002: ¥6,100,000 (Direct Cost: ¥6,100,000)
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| Keywords | Biological macromolecular assemblies / Synchrotron Radiation / Virus / Structure assembly of virus / X-ray crystal structure analysis / Infection mechanism of virus / Self-replication mechanism of virus / 多波長異常分散法 / タンパク質複合体 |
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| Research Abstract |
This project has been worked on the structural studies of Rice dwarf virus, which is a double-shelled virus with its molecular mass of about 7,500 kDa, to reveal its whole structure using x-ray crystallography, cryo-electron micrograph and molecular biological techniques.
Most of the structural studies have been done with the P2-deletion RDV particle, which contained all viral components except the P2 protein, where P2 protein had been removed from the sample during treatment with CCl_4 in the purification step. The 3.5Å resolution electron density map of RDV crystal, determined by x-ray crystallography, revealed almost complete structures of the inner- and outer-shells (P3 and P8 proteins) and a part of P7 protein. The interactions between the subunit proteins suggest how the 900 protein components are built into a higher-ordered virus core structure, and we proposed the model for the hierarchy of structural assembly. This model was strongly supported by some experiments using molecular biological techniques.
Internal structure of the virus particle was studied by combination of the single-particle analysis using cryo-electron micrograph and x-ray crystallography. Low-resolution cryo-EM map showed clear molecular envelop corresponding to the transcription complex (complex of P1,P5 and P7 proteins). The map also clearly showed at lease three layered-structure of dsRNA. Especially, dsRNAs in the first and second layer showed rod-shaped structure, which are packed into liquid crystal-like structure.
In addition, we have succeeded to purify the component proteins from the virus particle and/or from overexpressed sample, these samples were subjected to crystallization to reveal the high-resolution structure of the component proteins.
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