Evolution of Function of Bacterial Nucleoid-associated Proteins

该计划的重点是细菌类核的组织。细菌基因组与许多DNA结合蛋白相关。有些蛋白，如HU蛋白，是普遍存在的，而另一些蛋白仅由真细菌的一个子集编码。Dps（饥饿期间的DNA保护）蛋白属于后一类。一个主要目标是确定这些蛋白质家族中功能的分子决定因素。重点将放在耐辐射真杆菌Deinococcus radiodurans编码一个HU和两个Dps同系物。对D.对于耐辐射菌HU，将使用定点诱变和光化学交联来确定其在其优选底物（四向DNA连接）上的空间分布。在类核组织中的作用将通过在正常指数生长期间以及响应于DNA损伤的表达水平的体内定位和分析来评估。对于Dps-1，将在体外确定DNA相互作用的模式，并将确定其作为生长条件或环境应力的函数的表达。由于Dps-2被预测为非细胞质的，其细胞定位将被确定，沿着其响应于应激的DNA结合和调节模式。预计所提出的分析将不仅有助于我们对两个重要的类核蛋白家族成员的DNA结合机制的理解，而且有助于阐明细胞过程，这些细胞过程有助于对环境胁迫的特殊抗性，这是D.耐辐射的该计划的重点是通过为研究生和本科生提供获得研究经验的机会来整合研究和教学。将强调少数民族学生的参与，并期望各级学生在全国会议上传播其研究成果。

The focus of this program is on the organization of the bacterial nucleoid. The bacterial genome is associated with a number of DNA-binding proteins. Some, such as HU proteins, are ubiquitous, while others are only encoded by a subset of eubacteria. Dps (DNA protection during starvation) proteins belong to the latter category. A main goal is to define molecular determinants of function within these protein families. The focus will be on the radiation-resistant eubacterium Deinococcus radiodurans which encodes one HU and two Dps homologs. For D. radiodurans HU, its spatial disposition on its preferred substrate, the four-way DNA junction, will be determined using site-directed mutagenesis and photochemical crosslinking. Roles in nucleoid organization will be assessed by in vivo localization and analysis of expression levels during normal exponential growth as well as in response to DNA damage. For Dps-1, the mode of DNA interaction will be determined in vitro and its expression as a function of growth condition or environmental stress will be determined. As Dps-2 is predicted to be non-cytoplasmic, its cellular localization will be determined, along with its mode of DNA binding and regulation in response to stress. It is anticipated that the proposed analyses will contribute not only to our mechanistic understanding of DNA binding by members of two important families of nucleoid-associated proteins, but also to elucidating cellular processes that contribute to the exceptional resistance to environmental stress that is characteristic of D. radiodurans. This program focuses on integrating research and teaching by offering the opportunity for both graduate and undergraduate students to acquire research experience. The involvement of minority students will be emphasized, and students at all levels will be expected to disseminate their research at national meetings.