Structure and Mechanism of a Prion-remodeling Factor

描述（由申请人提供）：朊病毒是一种非常规的高感染性病原体，完全由一种采用异常构象的蛋白质组成。在哺乳动物中，朊病毒介导的感染是造成几种毁灭性的和总是致命的神经退行性疾病的原因，统称为传染性海绵状脑病。朊病毒疾病的标志是淀粉样蛋白的存在，其也与非朊病毒疾病的病理学相关，所述非朊病毒疾病的范围从阿尔茨海默病和亨廷顿病到系统性淀粉样变性。广泛而长期的研究目标是揭示分子伴侣在朊病毒复制中的功能作用。酵母为研究朊病毒复制机制提供了一个很好的范例。[PSI+]是增加无义密码子的翻译通读的酵母朊病毒。与哺乳动物朊病毒一样，[PSI+]完全由蛋白质组成，并由进化保守的翻译终止因子Sup 35 p（eRF 3）的自我复制淀粉样蛋白构象形成。[PSI+]的遗传和维持由Hsp 104控制，Hsp 104是一种600 kDa的、成环ATP依赖的蛋白质重塑机器，其在朊病毒复制和蛋白质解聚中与Hsp 70伴侣系统合作。本研究的目的是提供一个详细的机制，了解朊病毒重塑和蛋白解聚活动的热休克蛋白104及其细菌同源物ClpB。本研究的主要目的是：1）确定Hsp 104-底物复合物的三维结构; 2）研究Hsp 104与Hsp 70/Hsp 40之间的协同作用; 3）阐明Hsp 104双分子伴侣系统对蛋白质解聚和朊病毒复制的作用机制。为了解决我们的研究问题，我们将使用多方面的方法，包括混合结构生物学方法，蛋白质组学和化学生物学技术，以及酵母遗传学。这些方法的结合提供了一个强大的方法，以产生新的机制洞察到这个显着的家庭的ATP依赖的分子机器的结构-功能关系，以便这些信息可能被利用到工程师新的纳米机器与新的生物活性与潜在的应用在生物技术和纳米医学。

DESCRIPTION (provided by applicant): Prions are unconventional, highly infectious agents, which are composed entirely of a protein that adopts an abnormal conformation. In mammals, prion-mediated infections are responsible for several devastating and invariably fatal neurodegenerative diseases, collectively known as transmissible spongiform encephalopathies. A hallmark of prion diseases is the presence of amyloids, which are also associated with the pathology of non-prion diseases, ranging from Alzheimer's and Huntington's disease to systemic amyloidosis. The broad and long-term research objective is to uncover the functional role of molecular chaperones in prion replication. Yeast provides an excellent paradigm to investigate the mechanism of prion replication. [PSI+] is a yeast prion that increases translational read-through of nonsense codons. Like mammalian prions, [PSI+] consist entirely of protein and is formed by self-replicating amyloid conformers of the evolutionary conserved translation termination factor Sup35p (eRF3). The inheritance and maintenance of [PSI+] are governed by Hsp104, a 600-kDa, ring-forming ATP-dependent, protein-remodeling machine, which cooperates with the Hsp70 chaperone system in prion replication and protein disaggregation. The objective of this research is to provide a detailed mechanistic understanding of the prion-remodeling and protein disaggregating activities of Hsp104 and its bacterial homolog ClpB. Three specific aims are proposed: 1) to determine the 3D structure of an Hsp104-substrate complex, 2) to investigate the synergistic interaction between Hsp104 and Hsp70/Hsp40, and 3) to elucidate the mechanism of protein disaggregation and prion replication by the Hsp104 bi-chaperone system. To address our research questions, we will use a multi-facet approach consisting of hybrid structural biology methods, proteomic and chemical biology techniques, and yeast genetics. The combination of these methods provides a powerful approach to yield new mechanistic insight into the structure-function relationship of this remarkable family of ATP-dependent molecular machines in order that this information might be exploited to engineer new nano-machines with novel biological activities with potential applications in biotechnology and nano-medicine.