NMR Structure of the A-beta Peptide Inside Living Cells

活细胞内 A-β 肽的 NMR 结构

• 批准号: 9271867
• 负责人: MICHAEL Gerard ZAGORSKI
• 金额: $ 18.84万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-15 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9271867
• 关键词: Adopted; Alzheimer' s Disease; Amino Acids; Amyloid; Amyloid Fibrils; Amyloid beta-Protein; Amyloidosis; Area; Brain; Brain Diseases; Cell Nucleus; Cells; Chemicals; Complex; Coupled; Crowding; Data; Deposition; Detection; Development; Diffusion; Disease; Disease Progression; Drug Design; Environment; Escherichia coli; Eukaryotic Cell; Event; Goals; Human; In Vitro; Isotopes; Kinetics; Label; Laboratories; Lead; Lewy Bodies; Link; Measures; Methods; Mitochondria; Monitor; NMR Spectroscopy; Neurofibrillary Tangles; Nuclear Magnetic Resonance; Parkinson Disease; Pathogenicity; Patients; Peptides; Physiological; Principal Investigator; Process; Proteins; Relaxation; Reporting; Research; Resolution; Senile Plaques; Signal Transduction; Site; Solvents; Structure; Subcellular structure; Techniques; Transgenic Mice; Work; abeta accumulation; abeta deposition; alpha synuclein; amyloid formation; aqueous; base; beta pleated sheet; experience; experimental study; extracellular; flexibility; in vivo; inhibitor/antagonist; monomer; neuron loss; neurotoxic; neurotoxicity; novel; prevent; programs; protein structure; public health relevance; recombinant peptide; three dimensional structure; tool

 DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is characterized by the abundance of intraneuronal neurofibrillary tangles and the extracellular deposition of the Aβ peptide (hereafter referred to as Aβ) into amyloid. In AD brains, the Aβ undergoes a conversion (monomer → aggregate) and eventually precipitates as β-sheet like structures that have been linked to neuronal death. The chemical mechanisms associated with these conversions remains largely unknown. Due to its high propensity to aggregate, the Aβ is extremely difficult to handle in the laboratory. Despite these difficulties, our research group was one of the first to perform solution NMR studies of the Aβ, which along with subsequent work from other labs established that the Aβ adopts a predominantly random, unfolded structure in aqueous solution at neutral pH. A major advantage of the NMR approach is that it can provide atomic level aspects of the structures and dynamics in solution that are not available with other low-resolution techniques. In this application, we will take the next important and novel step, and will use NMR to study the structural ensembles of the Aβ peptide inside living eukaryotic cells. This work will establish whether or not the Aβ structures in vitro are the same as those in living cells in vivo. This is extremely important, in that the Aβ is an "intrinsically disordered" protein, which is a major clas of proteins that have a very flexible three- dimensional structure and are particularly sensitive t environmental perturbations, and that the complex, crowded environment of the cell could potentially alter the structure and function. We will perform two Specific Aims and attempt to determine the intracellular structure and dynamics in living cells using an array of NMR experiments. We will also explore the interactions between the Aβ and mitochondria. Furthermore, the classical view is that the Aβ is deposited extracellularly, although new data from transgenic mice and human patients demonstrates that the Aβ can likewise accumulate intraneuronally and contribute to AD progression, further emphasizing the importance of in-cell NMR studies.

 描述（由适用提供）：阿尔茨海默氏病（AD）的特征是神经内神经原纤维缠结的抽象和Aβ肽的细胞外沉积（以下称为Aβ）中的淀粉样蛋白。在AD大脑中，Aβ经历了转化率（单体→骨料），并最终作为与神经元死亡有关的β-折叠类似结构。与这些转化有关的化学机制在很大程度上仍然未知。由于其聚集的很高的承诺，在实验室中很难处理Aβ。尽管遇到了这些困难，但我们的研究小组是对Aβ进行溶液NMR研究的最早研究小组之一，该研究与其他实验室的后续工作一起确定，Aβ在中性pH下水溶液中采用了主要随机的，未展开的结构。 NMR方法的一个主要优点是，它可以提供解决方案中结构和动力学的原子水平方面，而其他低分辨率技术则无法提供。在此应用中，我们将采取下一个重要和新颖的步骤，并将使用NMR研究活着的真核细胞内Aβ肽的结构集合。这项工作将确定体外Aβ结构是否与体内活细胞中的Aβ结构相同。这非常重要，因为Aβ是一种“本质上无序”的蛋白质，它是具有非常灵活的三维结构并且特别敏感的环境扰动的主要蛋白质块，并且细胞的复杂，拥挤的环境可能会改变结构和功能。我们将执行两个特定的目的，并尝试使用NMR实验阵列来确定活细胞中细胞内结构和动力学。我们还将探索Aβ和线粒体之间的相互作用。此外，经典的观点是Aβ被细胞外沉积，尽管来自转基因小鼠和人类患者的新数据表明，Aβ同样可以积累内神经内神经性并有助于AD进展，进一步强调了细胞内NMR研究的重要性。