| Project/Area Number |
15370065
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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 | Tokyo Institute of Technology |
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Principal Investigator |
ARISAKA Fumio TOKYO INSTITUTE OF TECHNOLOGY, DEPT OF BIOMOLECULAR PROCESSING, ASSOCIATE PROFESSOR, 大学院・生命理工学研究科, 助教授 (80133768)
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| Co-Investigator(Kenkyū-buntansha) |
KANAMARU SHUJI TOKYO INSTITUTE OF TECHNOLOGY, DEPT OF BIOMOLECULAR PROCESSING, ASSISTANT PROFESSOR, 大学院・生命理工学研究科, 助手 (50376951)
TAKEDA SHIGEKI GUNMA UNIVERSITY, FAC ENGINEERING, ASSOCIATE PROFESSOR, 大学院・工学研究科, 助教授 (80282854)
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| Project Period (FY) |
2003 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥14,900,000 (Direct Cost: ¥14,900,000)
Fiscal Year 2005: ¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 2004: ¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2003: ¥9,200,000 (Direct Cost: ¥9,200,000)
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| Keywords | protein / bacteriophage / molecular machine / molecular recognition / molecular assembly / infection / analytical ultracentrifugation / X-ray crystallography / 微生物 / 超分子構造 / リゾチーム / テイルリゾチーム / 収縮姓尾部 / 感染機構 / バクテイオファージ / T4ファージ / プロセッシング / 蛋白質工学 / T4類縁ファージ / βヘリックス |
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| Research Abstract |
The purpose of the present study is to elucidate the mechanism of assembly and function of bacteriophage as a model system in terms of its molecular and atomic level. The major results we obtained were the following :
The most important progress in elucidating the mechanism of the structural transformation of the contractile tail of phage T4 came from the collaboration with Michael Rossmann's group at Purdue. Leiman in Rossmann's group succeeded in determining the low resolution structure of the contracted tail by 3D image reconstruction from cryo-EM. The fact that the same high resolution structure of the subunits can be fitted into the electron density of the contracted conformation indicates that the subunits themselves do not change their conformation significantly, but change the relative positions in the baseplate during conformational change of the baseplate. Using the extended and contracted images of the tail and interpolation, we were able obtain a reasonably detailed model of
… More
how the subunit rearrangement takes place. Gp11 strongly binds to the domain III and the N-terminus of gp12 which is the short tail fiber binds to domain IV of gp10 before structural change. The protein which plays a key role during the "hexagon" to "star" conformational change of the baseplate is gp10. Upon infection, as gp10 rotates around the longitudinal axis, gp11 which had been bound to gp12 now binds to gp34, the proximal long tail fiber, and the short tail fiber extends out from the bottom of the baseplate.
Secondly, we have succeeded in elucidating the tail lysozyme (gp5) structure, where the amino acid at 351^<st> position was replaced with seven different amino acids. One of them, S351L, of which the cleavage was completely suppressed, was expressed, purified and crystallized for X-ray diffraction. The structure of the S351L mutant showed that the cleavage site was highly exposed, as would be required for efficient digestion during phage maturation. Since the structure of gp5 does not resemble any known protease, the cleavage is most likely not autocatalytic, but the result of an E.coli protease. The mutant structure further demonstrates that the inhibition peptide from the neighboring subunit is already at the substrate recognition site prior to cleavage. Less
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