Structural Biology of Amyloid Beta-Protein

描述（由申请人提供）：我们假设淀粉样蛋白(A)组装成神经毒性低聚物和聚合物是阿尔茨海默病（AD）的一个重要神经发病过程。如果是这样，抑制组装或分离现有组装可能是有效的治疗方法。为了验证我们的假设，必须详细阐明A¿的结构生物学。什么构象形成低聚物？通过什么机制？这样形成的低聚物的结构是什么？每种低聚物的相对毒性是什么？许多人，包括我们自己，都在努力将结构与生物活性的测量联系起来。最近的研究表明，二聚体或三聚体组合是重要的神经毒素，但六聚体、非三聚体、十二聚体和更大的低聚体也被证明是有效的神经毒素。本提案的长期目标是将过去简单的四级结构确定转移到阐明A¿单体的二级和三级结构动力学和确定单体寡聚的机制。这意味着最终理解控制动力学的原子间相互作用，并在原子分辨率上确定治疗靶点。这种“以知识为基础”的方法不同于高通量筛选策略，但它是一种补充。这两种方法都应该执行，以最大限度地确定有效的，改善疾病的治疗剂的机会。在此，我们提出：(1)阐明A¿单体折叠和自组装的物理生化；(2)建立由此形成的A¿组装体的结构-神经毒性关系。为此，我们将化学合成A -肽，其中特定的氨基酸和化学键被改变，然后研究这些肽的构象动力学和组装。这些改变的位置，以及改变本身，都是经过仔细选择的，以便揭示A¿分子的关键结构特征，这些特征控制着A¿分子组装成破坏或杀死神经元的结构。我们将鉴定、分离和结构表征特定类型的组装，然后通过处理培养中的原代神经元定量确定每种组装的毒性活性。在第一个目标中获得的组件结构的理解深度将是前所未有的。因此，通过这种“结构-活性关联”过程获得的知识有望提供最准确的评估，以确定哪些组合，以及这些组合上的哪些结构（在原子分辨率上）应该用于靶向治疗。除了有助于提高对阿尔茨海默病及其治疗的理解外，拟议项目的结果应该与其他与异常蛋白质组装相关的神经退行性疾病的研究相关。这些疾病包括帕金森氏症、亨廷顿氏症、肌萎缩性侧索硬化症、家族性淀粉样多发性神经病和朊病毒。

DESCRIPTION (provided by applicant): We hypothesize that amyloid ¿-protein (A¿) assembly into neurotoxic oligomers and polymers is a seminal neuropathogenetic process in Alzheimer's disease (AD). If so, assembly inhibition or dissociation of existing assemblies could be effective therapeutic approaches. To test our hypothesis, the structural biology of A¿ must be elucidated in detail. What conformers form oligomers? By what mechanism? What are the structures of the oligomers thus formed? What is the relative toxicity of each oligomer species? Many, including ourselves, have striven to correlate structure with measures of biological activity. Recent work has suggested that dimeric or trimeric assemblies are important neurotoxins, but hexameric, nonameric, dodecameric, and larger oligomers also have been shown to be potent neurotoxins. The long-term goal of this proposal is to move past simple quaternary structure determination to elucidation of A¿ monomer secondary and tertiary structure dynamics and the determination of mechanisms of monomer oligomerization. This means eventually understanding the interatomic interactions that control the dynamics, and in doing so, identifying therapeutic targets at atomic resolution. This "knowledge-based" approach is distinct from, but complementary to, high-throughput screening strategies. Both approaches should be executed to maximize the chances for identifying efficacious, disease-modifying therapeutic agents. We propose here to: (1) elucidate the physical biochemistry of A¿ monomer folding and self-assembly; and (2) establish structure-neurotoxicity relationships of the A¿ assemblies thus formed. To do so, we will chemically synthesize A¿ peptides in which specific amino acids and chemical bonds are altered and then study the conformational dynamics and assembly of these peptides. The positions of these alterations, and the alterations themselves, have been chosen carefully so as to reveal the key structural features of the A¿ molecule that control its assembly into structures that damage or kill neurons. We will identify, isolate, and structurally characterize specific types of assemblies and then determine quantitatively the toxic activity of each assembly by treating primary neurons in culture. The depth of understanding of the structures of the assemblies obtained in the first aim will be unprecedented. Thus the knowledge gained through this "structure-activity correlation" process is expected to provide the most accurate assessment of which assemblies, and which structures (at atomic resolution) on these assemblies, should be targeted therapeutically. In addition to its contributions to an improved understanding of AD and its treatment, results of the proposed project should have relevance for studies of other neurodegenerative diseases linked to aberrant protein assembly. These include Parkinson's, Huntington's, amyotrophic lateral sclerosis, familial amyloid polyneuropathy, and the prionoses. PUBLIC HEALTH RELEVANCE: This project will advance our understanding of how a protein, the amyloid-protein, causes Alzheimers disease. This understanding can be translated directly into the development of a new class of drugs that have the potential to modify or cure the disease. The project also will be of relevance to Parkinsons, Huntingtons, amyotrophic lateral sclerosis, familial amyloid polyneuropathy, the prionoses, and other neurodegenerative diseases of aging.