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Role of three dimensional structure of proteins in signal transduction

蛋白质三维结构在信号转导中的作用

基本信息

批准号：
06276105
负责人：
INAGAKI Fuyuhiko
金额：
$ 68.03万
依托单位：
Tokyo Metropolitan Organization for Medical Research
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1997
项目状态：
已结题

项目摘要

The aim of the present research was to elucidate the roles of proteins in signal transduction process on the structural basis. We also developed the protein expression and purification systems for preparation of amount of proteins required for structural studies. In the course of the present research project, a number of protein structures were determined by either NMR spectroscopy or X-ray crystallography. Grb2 n-SH3 and SH2 domains complexed with proline rich peptide from Sos and phosphotyrosine containing peptide from Shc, respectively were determined and the general mechanism for recognition of target sequences were elucidated. Three dimensional structure of destrin was determined by NMR. Although the sequence homology between destrin family and gelsolin family proteins was not present, the three dimensional structure of both proteins were very similar, suggesting the same mechanism for actin severing. Thus, the general mechanism for severing of the actin fiber was proposed. The three dimensional structure of midkine was also determined by NMR and elucidated the mechanism of dimer formation on heparin oligosaccharides. The dimer formation of midkine seems to be essential for activation of the receptor and its biological function. We also determined the CRD domain of PKC and suggested the mechanism of DAG recognition. By X-ray crystallography, a number of protein structures were determined including transmitter and receiver domains of bacterial signal transduction system. We also made protein expression and purification system for N-WASP, Tob and CAF1. N-WASP is essential for formation of philopodia in response to the growth factor receptor signal. Tob and CAF1 play pivotal role in cell cycling. The structural studies started for both proteins.

本研究的目的是在结构基础上阐明蛋白质在信号转导过程中的作用。我们还开发了蛋白质表达和纯化系统，用于制备结构研究所需的大量蛋白质。在本研究项目的过程中，通过核磁共振光谱或x射线晶体学确定了许多蛋白质结构。确定了Grb2 n-SH3和SH2结构域分别与来自Sos的富含脯氨酸肽和来自Shc的含磷酸酪氨酸肽络合，并阐明了识别靶序列的一般机制。用核磁共振法测定了样品的三维结构。尽管destrin家族蛋白与gelsolin家族蛋白的序列不存在同源性，但两者的三维结构非常相似，提示肌动蛋白断裂的机制相同。因此，提出了肌动蛋白纤维断裂的一般机制。通过核磁共振测定了midkine的三维结构，并阐明了其在肝素寡糖上形成二聚体的机理。midkine的二聚体形成似乎是受体激活及其生物学功能所必需的。我们还确定了PKC的CRD结构域，并提出了DAG识别的机制。通过x射线晶体学，确定了细菌信号转导系统的一些蛋白质结构，包括传递域和接收域。我们还制作了N-WASP、Tob和CAF1蛋白的表达纯化系统。N-WASP是响应生长因子受体信号形成philopodia所必需的。Tob和CAF1在细胞周期中起关键作用。对这两种蛋白质的结构研究开始了。