Structural and Functional Studies of Hsp70 Molecular Chaperones

Hsp70 分子伴侣的结构和功能研究

• 批准号: 9067389
• 负责人: Qinglian Liu
• 金额: $ 28.98万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9067389
• 关键词: ATP phosphohydrolase; Accounting; Allosteric Regulation; Alzheimer' s Disease; Binding; Binding Sites; Biochemical; Biochemistry; Color; Coupling; Disease; Escherichia coli; Genetic; Goals; Health; Heart; Homeostasis; Human; Kinetics; Length; Life; Link; Malignant Neoplasms; Manuscripts; Membrane; Modeling; Molecular; Molecular Chaperones; Nature; Neurodegenerative Disorders; Nucleotides; Organism; P2 peptide; Parkinson Disease; Peptide Conformation; Peptides; Play; Property; Proteins; Research; Resolution; Role; Side; Site; Structure; Substrate Domain; Testing; Therapeutic Intervention; Transportation; X-Ray Crystallography; Yeasts; base; computational chemistry; design; human disease; innovation; insight; interdisciplinary approach; novel; novel therapeutics; polypeptide; protein folding; unpublished works

DESCRIPTION (provided by applicant): Hsp70s are ubiquitous and highly conserved molecular chaperones that play multiple essential roles in maintaining cellular protein homeostasis through assisting in protein folding, assembly, degradation, and transportation across membrane. The fundamental importance of maintaining protein homeostasis inevitably links Hsp70s with many destructive human diseases, most notably cancers and neurodegenerative disorders, such as Parkinson's and Alzheimer's diseases. Thus, elucidating the structural and biochemical properties of Hsp70s will not only advance our understanding of the basic molecular mechanism of Hsp70-assited protein folding, but also provide crucial insights regarding how to target Hsp70s for therapeutic interventions in treating cancers and neurodegenerative disorders. Hsp70s have three key biochemical activities that are at the heart of the chaperone activity: ATPase, peptide substrate binding, and ATP-induced allosteric coupling. In spite of extensive efforts, the very basic mechanism of Hsp70-assisted protein folding is still ill-defined due to the lack of in-depth understanding o two key biochemical activities: 1) ATP-induced allosteric coupling is central to Hsp70s' chaperone activity~ however, all previous studies had failed to reveal the molecular mechanism. 2) The well-established single peptide binding site on Hsp70s has made it difficult to explain the efficient chaperone activity. Could there be additional peptide binding sites on Hsp70s that account for the high efficiency? Thus, the overall objective of this proposal is to analyze these two key biochemical activities in order to dissect the basic mechanisms of Hsp70 chaperone function. Recently, we solved the first crystal structure of an intact Hsp70 in the allosteric active state, and discovered a novel peptide substrate binding site on Hsp70s. Based on these original discoveries, we propose the following two Specific Aims: 1) elucidate the molecular mechanism of the ATP-driven allosteric coupling in Hsp70s, and 2) characterize a novel peptide binding site on Hsp70s and investigate its role in protein folding. To achieve our goal, we use a multidisciplinary approach combining X-ray crystallography, biochemistry, NMR, EPR, computational chemistry, and yeast and E.coli genetics. We expect that successful completion of this proposal will help us realize our long-term goal, which is to establish a thoroug mechanism understanding of the very basic mechanism of Hsp70 chaperone activity in protein folding.

描述（由申请人提供）：HSP70是普遍存在且高度保守的分子伴侣，在维持细胞蛋白质稳态方面起着多种基本作用，通过协助蛋白质折叠，组装，降解，降解和跨膜的运输。 维持蛋白质稳态的基本重要性不可避免地将HSP70与许多破坏性的人类疾病联系起来，最著名的是癌症和神经退行性疾病，例如帕金森氏症和阿尔茨海默氏病。因此，阐明HSP70S的结构和生化特性不仅会提高我们对HSP70蛋白质折叠的基本分子机制的理解，而且还提供了有关如何针对HSP70来治疗治疗癌症和神经变性性分离的治疗干预措施的关键见解。 HSP70具有三种关键的生化活性，这是伴侣活性的核心：ATPase，肽底物结合和ATP诱导的变构耦合。 尽管做出了广泛的努力，但由于缺乏深入了解HSP70辅助蛋白折叠的非常基本的机制仍然不明显，o两种关键的生物化学活动：1）ATP诱导的变构耦合对HSP70S的伴侣的伴侣的核心核心是HSP70S的伴侣活性〜但是，所有先前的研究都未能揭示出分子机械性的。 2）HSP70S上建立良好的单肽结合位点使得难以解释有效的伴侣活性。 HSP70S上是否有其他肽的结合位点可以说明高效率？ 因此，该提案的总体目的是分析这两种关键的生化活动，以剖析HSP70伴侣功能的基本机制。 最近，我们在变构活性状态下解决了完整HSP70的第一个晶体结构，并发现了一种新颖 HSP70S上的肽底物结合位点。基于这些原始发现，我们提出了以下两个特定目的：1）阐明了HSP70中ATP驱动的变构耦合的分子机制，以及2）表征HSP70上新型肽结合位点并研究其在蛋白质折叠中的作用。 为了实现我们的目标，我们使用X射线晶体学，生物化学，NMR，EPR，计算化学以及酵母和大肠杆菌遗传学的多学科方法。我们预计该提案的成功完成将有助于我们实现我们的长期目标，即建立对蛋白质折叠中HSP70伴侣活性非常基本机制的胸腔机制理解。