Structure/Mechanism of a Prion-remodeling Factor

描述（由申请人提供）：朊病毒是高度感染性的蛋白质类物质，可引起多种毁灭性和致命性神经退行性疾病。例如人类的克雅氏病和库鲁病、牛的牛海绵状脑病以及麋鹿和鹿的慢性消耗性疾病。我们研究的广泛和长期目标是进一步了解朊病毒和朊病毒疾病的分子。 [PSI+]是增加无义密码子的翻译通读的酵母朊病毒。像哺乳动物的朊病毒一样，酵母朊病毒完全由蛋白质组成。[PSI+]由Sup 35的自我复制淀粉样蛋白构象形成，其最终导致淀粉样蛋白原纤维的形成，这是朊病毒疾病的标志。最有趣的是，[PSI+]的遗传、繁殖和消除由Hsp 104分子伴侣控制。热休克蛋白104是一种ATP依赖性蛋白质重塑因子，它首先被发现是酵母应激反应中的一种重要成分，可以将应激损伤的蛋白质从先前的聚集状态中拯救出来。虽然解聚蛋白质的能力与细菌ClpB共享，但ClpB既不促进繁殖也不促进[PSI+]的消除，这表明朊病毒重塑和蛋白质解聚活性在机制上是不同的。 该建议的目的是提供一个详细的机制的理解，热休克蛋白104在prionogenesis的作用。提出了以下具体目标：（1）确定Hsp 104的高分辨率晶体结构，（2）剖析Hsp 104朊病毒重塑活性的生化机制，和（3）确定Hsp 104/Sup 35复合物的3D结构。为了实现我们的目标，我们将结合联合收割机三维结构的研究与生物化学和高通量蛋白质组学的方法。这些技术的结合将为朊病毒重塑机制提供实质性的新见解，朊病毒重塑是Hsp 104特有的活性。此外，我们提出的研究也可能提供为什么分子伴侣不能预防朊病毒感染的答案，并可能在寻找朊病毒感染和其他蛋白质错误折叠疾病的潜在治疗方法中开辟新的途径。 朊病毒是一种非传统的、高度传染性的病原体，可引起几种毁灭性的、总是致命的神经退行性疾病，统称为传染性海绵状脑病。分子伴侣提供了对抗朊病毒感染和其他蛋白质错误折叠疾病的第一道防线。本文主要探讨热休克蛋白104的结构和作用机制。热休克蛋白104是一种朊病毒重塑因子，也是应激反应的重要分子伴侣。

DESCRIPTION (provided by applicant): Prions are highly infectious proteinaceous agents responsible for several devastating and invariably fatal neurodegenerative diseases. Examples include Creutzfeldt-Jacob disease and kuru in humans, bovine spongiform encephalopathy in cattle, and chronic wasting diseases in elk and deer. The broad and long-term objective of our research is to further our molecular understanding of prions and prion diseases. [PSI+] is a yeast prion that increases translational read-through of nonsense codons. Like mammalian prions, yeast prions consist entirely of protein. [PSI+] is formed by self-replicating amyloid conformers of Sup35, which ultimately result in the formation of amyloid fibrils, a hallmark of prion diseases. Most interestingly, the inheritance, propagation, and elimination of [PSI+] are governed by the Hsp104 molecular chaperone. First discovered as an essential component in the yeast stress response, Hsp104 is an ATP-dependent protein- remodeling factor, which can rescue stress-damaged proteins from a previously aggregated state. While the ability to disaggregate proteins is shared with bacterial ClpB, ClpB neither promotes propagation nor facilitates elimination of [PSI+], suggesting that prion-remodeling and protein-disaggregating activities are mechanistically distinct. The goal of this proposal is to provide a detailed mechanistic understanding of the role of Hsp104 in prionogenesis. The following specific aims are proposed: (1) to determine the high-resolution crystal structure of Hsp104, (2) to dissect the biochemical mechanism of Hsp104 prion-remodeling activity, and (3) to determine the 3D structure of an Hsp104/Sup35 complex. To achieve our goals, we will combine three-dimensional structural studies with biochemical and high-throughput proteomic approaches. The combination of these techniques will provide substantial new insight into the mechanism of prion remodeling, an activity unique to Hsp104. Moreover, our proposed studies may also provide answers to why molecular chaperones cannot prevent prion infections, and may open new avenues in the search for a potential treatment of prion infections and other protein misfolding diseases. Prions are unconventional, highly infectious agents, responsible for several devastating and invariably fatal neurodegenerative diseases collectively known as transmissible spongiform encephalopathies. Molecular chaperones provide the first line of defense against prion infections and other protein misfolding diseases. Here we will investigate the structure and mechanism of Hsp104, a prion-remodeling factor and essential molecular chaperone of the stress response.