Structure and Function of Nucleosome Assembly Protein 1

描述（由申请人提供）：染色质起着两个重要的，但本质上是相互排斥的作用。一方面，染色质负责极端程度的DNA压实，这使得真核生物基因组能够适应细胞核的范围。另一方面，它必须允许DNA的局部分解，以允许细胞机制进入基因组。介导不同致密染色质状态之间相互转化的活动是所有需要获得基因组DNA的过程的关键调节因子。对这些生物学相关过程的机制和结构的了解是有限的。本文将研究组蛋白伴侣的核小体组装蛋白1 （NAP1）家族在维持和调节染色质结构和流动性中的作用。目的1和2利用x射线衍射以及生化和体内方法解决伴侣蛋白与组蛋白相互作用的结构和分子基础。nap1介导的组蛋白去除和组蛋白交换的机制和生物学作用将通过严格的动力学分析结合体内染色质结构检查来研究（目的3）。最后，在第4篇论文中，我们将对翻译后修饰调节后生动物中NAP1活性的新假设进行验证。该建议的优势在于结合了广泛的结构，生物物理，生化和遗传方法，以解决如何使细胞机器获得紧密DNA这一高度重要的问题。

DESCRIPTION (provided by applicant): Chromatin plays two essential, yet by their very nature mutually exclusive roles. On one hand, chromatin is responsible for the extreme degree of DNA compaction that allows the eukaryotic genome to fit into the confines of the nucleus. On the other hand, it must permit the local unraveling of DNA to grant access of the cellular machinery to the genome. Activities that mediate the inter- conversion between various levels of compacted chromatin states are key regulators of all processes requiring access to genomic DNA. Mechanistic and structural insight into these biologically relevant processes is limited. Here, the role of the nucleosome assembly protein 1 (NAP1) family of histone chaperones in maintaining and modulating chromatin structure and fluidity will be investigated. Aims 1 and 2 address the structural and molecular basis for chaperone interaction with histones, using x-ray diffraction as well as biochemical and in vivo approaches. The mechanism and biological role of NAP1-mediated histone removal and histone exchange will be studied by rigorous kinetic analysis combined with in vivo examination of chromatin structure (aim 3). Finally, the novel hypothesis that a posttranslational modification regulates NAP1 activity in metazoans will be put to test in aim 4. The strength of this proposal lies in a combination of a broad spectrum of structural, biophysical, biochemical and genetic approaches to address the highly significant question of how compacted DNA is made available to the cellular machinery. The molecular mechanism and biological relevance of histone chaperone-mediated chromatin structure modulation will be investigated, using a combination of structural, biophysical, and genetic approaches.