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Architecture and Principle of Formation of the Baseplate of Bacteriophage T4

噬菌体T4底板的结构和形成原理

基本信息

批准号：
10480178
负责人：
ARISAKA Fumio
金额：
$ 4.93万
依托单位：
Tokyo Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

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.

尾溶菌酶gp5是大肠杆菌的结构必需组分，具有溶菌酶活性，可在感染时局部消化宿主大肠杆菌的肽聚糖，使尾管到达内膜。gp5前体的切割发生在Ser351和Ala352之间。然而，C末端结构域作为复合体的一部分，只有在感染时才会分离。我们已经证明c端结构域存在于尾巴和病毒粒子中。研究还表明，感染细胞的尾部也保留了c端结构域。最近，普渡大学Michael G.Rossmann教授实验室的博士后Shuji kanamaruu成功地确定了gp5-gp27杂六聚体晶体衍射的相位。这种复合物的结构现在正在被解开，但它已经揭示了使gp5成为三聚体的细长的c端三重-螺旋。本课题要解决的另一个结构是底板楔体的起始配合物gp10-gp11配合物。认为它构成六角形底板顶点处的尾销，从底板底部伸出。通过SDS-PAGE、Edman降解和超离心分析表明，该配合物为异六聚体，亚基组成为（gp10）_3(gp11)_3。最近获得了适合x射线分析的配合物晶体，等待进行物相测定。作为结晶的第三个靶蛋白，尾部完成蛋白gp15也被过表达结晶。它形成一个同相六聚体，并可能形成管环的最后一个环。人们正在努力获得适合x射线衍射的更大的晶体。