| Project/Area Number |
10480178
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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 Grad. School of Biosci. & Bioeng. Assoc. Professor, 大学院・生命理工学研究科, 助教授 (80133768)
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| Project Period (FY) |
1998 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥7,700,000 (Direct Cost: ¥7,700,000)
Fiscal Year 2000: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 1999: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1998: ¥4,200,000 (Direct Cost: ¥4,200,000)
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| Keywords | molecular chaperone / association equilibrium / sedimentation equilibrium / thermal denaturation / Gdn-HCl denaturation / tail lysozyme / infection / X-ray crystallography / バクテリオファージ / プロセッシング / 分子集合 / X線結晶解析 / four helix-bumdie / 偽復帰突然変異体 |
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| Research Abstract |
Tail lysozyme, gp5, is a structurally essential component and possesses lysozyme activity which locally digests peptidoglycan of the host E.coli upon infection so that the tail tube can reach the inner membrane. Cleavage of gp5 precursor takes place between Ser351 and Ala352 during processing. The C- terminal domain, however, stays as part of the complex and will be detached only upon infection. We have demonstrated that the C-terminal domain is present in both tails and the virion. It was also shown that the tails in infedted cells also retain the C-terminal domain. Very recently, Shuji Kanamaruwho is a post-doctoral fellow at Purdue in Prof. Michael G.Rossmann's lab succeeded in determining the phase of diffractions from the crystal of gp5-gp27 hetero-hexamer. The structure of the complex is now being solved, but it already revealed the slender C-terminal triple beta-helix that makes gp5 a trimer. Another structure to be solved in this project is the initiation complex of the wedge of the baseplate, gp10-gp11 complex. It is considered to constitute the tail pins at the vertices of the hexagonal baseplate, which protrude from the bottom of the baseplate. Based on SDS-PAGE and Edman degradation of the complex and also analytical ultracentrifugation, it was shown that the complex is a hetero-hexamer with the subunit composition of (gp10)_3(gp11)_3. Crystal of the complex suitable for X-ray analysis has been recently been obtained which awaits the phase determination. As the third target protein for crystallization, gp15, the tail completion protein, was also overexpressed and crystallized. It forms a homo-hexamer and it is likely to form the last annulus of the tube ring. Efforts is being made to obtain larger crystals suitable for X-ray diffraction.
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