Extending the limits of cryo-EM to better understand TTR misfolding and aggregation

扩展冷冻电镜的局限性以更好地了解 TTR 错误折叠和聚集

• 批准号: 10263946
• 负责人: Gabriel C Lander
• 金额: $ 26.63万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10263946
• 关键词: 3-Dimensional; Adopted; Affect; Age; Alzheimer' s Disease; Alzheimer' s disease patient; Amyloid; Amyloid Fibrils; Amyloid beta-Protein; Amyloidosis; Architecture; Binding; Binding Proteins; Biochemical; Cardiomyopathies; Carrier Proteins; Cells; Cerebrospinal Fluid; Complex; Congestive Heart Failure; Cryoelectron Microscopy; Crystallization; Degenerative Disorder; Development; Diastolic heart failure; Disease; Disease Progression; Dissociation; Drug Design; Event; Funding; Future; Goals; Grant; Hepatocyte; Imaging technology; Individual; Kinetics; Lead; Life Expectancy; Ligand Binding; Ligands; Light; Link; Methodology; Methods; Mitotic; Molecular; Molecular Conformation; Molecular Probes; Names; Pathogenesis; Pathologic; Pathology; Pathway interactions; Patients; Peptides; Pharmaceutical Preparations; Plant Roots; Population; Prealbumin; Prevalence; Process; Property; Protein Export Pathway; Proteins; Protomer; Resolution; Restrictive Cardiomyopathy; Role; Serum; Structure; Therapeutic; Therapeutic Intervention; Thyroxine; Time; Tissues; Work; X-Ray Crystallography; age related; aggregation pathway; amyloidogenesis; base; combat; cytotoxic; design; fibrillogenesis; image processing; improved; insight; monomer; network dysfunction; non-Native; older patient; particle; prevent; protein aggregation; protein folding; protein misfolding; proteostasis; proteotoxicity; small molecule; therapeutic development; three dimensional structure

Appl ID: 9981223 Grant Number : 1R21 AG067594-01 PI Name : Lander, Gabriel Grant Title : Extending the limits of cryo-EM to better understand TTR misfolding and aggregation PROJECT SUMMARY The conversion of natively folded proteins into non-functional aggregates is associated with a wide range of age-related degenerative diseases, including Alzheimer’s Disease (AD). Of great concern is the prediction that these diseases will become more prevalent in prevalence as our nation’s life expectancy increases. The development of therapeutics to prevent or reverse the protein misfolding events implicated in degenerative diseases has thus become the focus of many investigative efforts. The protein transthyretin (TTR), a thyroxine and holoretinol carrier protein exported to cerebrospinal fluid (CSF) and serum, is one such protein that demonstrates increased propensity to adopt a non-native fold and form insoluble aggregates with age. This process can occur in the native, wild type form of the TTR protein, and is responsible for wild type TTR amyloidosis (also known as senile systemic amyloidosis), which causes restrictive cardiomyopathy. Notably, there is evidence that TTR interacts with the Aβ peptide, thereby preventing Aβ fibril formation and aggregation. Studies demonstrating that the CSF of AD patients contain substantially lower concentrations of TTR than in the CSF of age-matched non-AD individuals supports a neuroprotective role of TTR. We posit that age-related TTR misfolding and aggregation abolishes the capacity of TTR to prevent Aβ fibril formation and the subsequent onset of AD. In order to better understand how a natively folded wild type TTR protein becomes predisposed to misfolding events, we seek funding to develop structural approaches to study both the native and aberrant forms of TTR. Detailed structural information regarding the destabilization and non-native oligomerization of this protein will profoundly impact our understanding of TTR misfolding and fibrillogenesis, and could lead to the development of more potent TTR stabilizing drugs that could restore neuroprotective properties of TTR in AD patients. The first aim we will push the limits of size and resolution attainable by cryo-electron microscopy to examine the high-resolution three-dimensional structures of destabilized TTR tetramers and compare them to tetramers that are stabilized by small molecule ligands, revealing how the incorporation of misfolded subunits or the improper incorporation of natively folded subunits impact TTR stability at an atomic level. The second aim will define the architecture of TTR aggregates with atomic precision in order to shed light on the assembly pathways and how different oligomeric states differentially contribute to a variety of distinct pathogeneses. Notably, since perturbations of the native folding pathway of wild type proteins are the likely cytotoxic drivers of AD, the findings of this work will likely have far-reaching impact beyond TTR amyloidoses.

应用ID：9981223 赠款编号：1R21 AG067594-01 PI名称：Lander，Gabriel 赠款标题：扩大低温EM的限制以更好地了解TTR MIDDOLLED和聚合 项目摘要 本质上折叠的蛋白质转化为非功能骨料与包括年龄相关的变性疾病（包括阿尔茨海默氏病）（AD）有关。人们非常关心的是，随着我们国家的预期寿命增加，这些疾病将在流行率上变得更加普遍。因此，在退化性疾病中实施的蛋白质错误折叠事件的理论发展已成为许多调查工作的重点。导出至脑脊液（CSF）和血清的蛋白质经甲状腺素（TTR）是一种蛋白质，是一种这样的蛋白质，表现出一种越来越多的蛋白质，它表现出增加非本质折叠并形成不溶性折叠的诺言。该过程可以以TTR蛋白的天然野生型形式发生，并导致野生型TTR淀粉样变性（也称为老年系统性淀粉样变性），这会导致限制性心肌病。值得注意的是，有证据表明TTR与Aβ肽相互作用，从而防止Aβ原纤维形成和聚集。研究表明AD的CSF 患者的TTR浓度大大低于年龄匹配的非AD个体的CSF，支持TTR的神经保护作用。我们将与年龄相关的TTR杂交和聚集取消了TTR的能力防止Aβ原纤维形成和随后的AD发作。为了更好地理解本质上折叠的野生型TTR蛋白如何易于折叠事件，我们寻求资金来开发结构方法研究TTR的天然和异常形式。有关该蛋白质不稳定和非本地寡聚化的详细结构信息将对我们对TTR错误折叠和原纤维发生的理解产生深远的影响，并可能导致可能恢复AD患者TTR神经保护性质的更多潜在TTR稳定药物的发展。 The first aim we will push the limits of size and resolution attainable by cryo-electron microscopy to exam the High-resolution three-dimensional structures of destabilized TTR tetramers and compare them to tetramers that are stabilized by small molecule ligands, revealing how the incorporated influence of misfolded subunits or the improper incorporated impact TTR stability at an atomic level.第二个目的将用原子精度定义TTR聚集体的结构，以阐明组件途径以及不同的低聚物状态如何对各种不同的病原体有所不同。尤其， 由于野生型蛋白的天然折叠途径的扰动是AD的可能的细胞毒性驱动因素，因此这项工作的发现可能会在TTR淀粉样蛋白以外具有深远的影响。