Copper-Induced Amyloidosis Studied by Mass Spectrometry

通过质谱研究铜诱导的淀粉样变性

• 批准号: 7458170
• 负责人: RICHARD W VACHET
• 金额: $ 21.73万
• 依托单位: UNIVERSITY OF MASSACHUSETTS AMHERST
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2009-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7458170
• 关键词: Alzheimer' s Disease; Amino Acids; Amyloid; Amyloid Fibrils; Amyloidosis; Binding; Binding Proteins; Binding Sites; Condition; Copper; Detection; Development; Dialysis procedure; Disease; Evolution; Human; Label; Lead; Mass Spectrum Analysis; Measures; Metals; Methods; Modeling; Molecular; N-terminal; Nature; Non-Insulin-Dependent Diabetes Mellitus; Outcomes Research; Parkinson Disease; Peroxonitrite; Physical Dialysis; Play; Principal Investigator; Process; Protein Binding; Protein Footprinting; Proteins; Range; Reaction; Reagent; Relative (related person); Resolution; Role; Solvents; Spectrometry; Structure; System; Techniques; Therapeutic; Time; amyloid fibril formation; amyloid formation; base; design; fibrillogenesis; human disease; in vivo; microwave electromagnetic radiation; novel strategies; numb protein; oxidation; programs; tool

DESCRIPTION (provided by applicant): An increasing number of proteins are known to form amyloid fibrils in vivo, and the formation of these fibrils is implicated in several diseases. One of these proteins, human (B-2-microglobulin (B2m), can form amyloid fibrils in the presence of Cu (ll), and these fibrils are presumed to be the main pathogenic process underlying dialysis-related amyloidosis (DRA). Like other amyloid systems, p2m fibril formation proceeds by partial protein unfolding, subsequent oligomerization, and eventual elongation to form mature fibrils. While aspects of general amyloid formation are understood, molecular-level information is lacking for most amyloid systems; however, this information is critical for the rational design of therapeutics against amyloid diseases like DRA. We plan to obtain amino acid-level information of the unfolding and oligomerization process associated with B2m fibrillogenesis. To do so, we will apply, optimize, and develop 3 mass spectrometry (MS)-based tools with the necessary temporal and spatial resolution. (1). Metal-catalyzed oxidation (MCO) reactions with MS detection will provide information about the evolution of Cu (ll)-B2m interactions during unfolding and oligomerization. Improvements will also be made to the MCO/MS method in order to simplify the analysis and increase the temporal resolution of this technique. (2). A new 'detuned' MCO/MS method will measure the tertiary structural changes that occur around Cu (ll) during (B2m unfolding and oligomerization. The scope of this new method will be studied, including its potential to provide information about the relative distances of amino acids from Cu (ll). (3). Radical-based protein footprinting will be used to study changes in B2m structure during unfolding and oligomerization. Peroxynitrite will be investigated as a new protein footprinting reagent. We hypothesize that Cu (ll) binding destabilizes the N-terminal region of B2m. This destabilization leads to partial unfolding, and the subsequent structural changes that lead to oligomerization will be elucidated using our 3 complementary structural analysis tools.

描述(申请人提供)：越来越多的蛋白质在体内形成淀粉样纤维，这些纤维的形成与几种疾病有关。其中，人(B-2-微球蛋白，B2M)可在铜离子存在下形成淀粉样纤维，这些纤维被认为是透析相关淀粉样变性(DRA)的主要致病过程。与其他淀粉样蛋白系统一样，p2m原纤维的形成是通过部分蛋白质的展开、随后的寡聚和最终的伸长形成成熟的原纤维。虽然人们已经了解了淀粉样蛋白形成的各个方面，但大多数淀粉样蛋白系统缺乏分子水平的信息；然而，这些信息对于合理设计针对DRA等淀粉样蛋白疾病的治疗方案至关重要。我们计划获得与B2M纤维蛋白形成相关的去折叠和寡聚过程的氨基酸水平信息。为此，我们将应用、优化和开发具有必要时间和空间分辨率的3种基于质谱学(MS)的工具。(1)。金属催化氧化(MCO)反应与MS检测将提供有关铜(11)-B2M相互作用在去折叠和齐聚过程中的演变的信息。还将对MCO/MS方法进行改进，以简化分析并提高该技术的时间分辨率。(2)。一种新的“失谐”MCO/MS方法将测量在B2M展开和齐聚过程中发生在Cu(11)周围的三级结构变化。将研究这种新方法的范围，包括它提供有关氨基酸与铜(11)的相对距离的信息的潜力。(3)。基于自由基的蛋白质足迹将被用来研究B2M结构在去折叠和寡聚过程中的变化。过氧亚硝酸盐将作为一种新的蛋白质足迹试剂进行研究。我们假设铜(11)结合破坏了B2M的N-末端区域的稳定性。这种不稳定会导致部分展开，随后导致齐聚的结构变化将使用我们的3个互补结构分析工具来阐明。