Detection and structural analysis of misfolded proteins causing neurodegeneration

引起神经变性的错误折叠蛋白的检测和结构分析

基本信息

批准号：
8286425
负责人：
DAVID W COLBY
金额：
$ 20.8万
依托单位：
UNIVERSITY OF DELAWARE
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-01-01 至 2013-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8286425
关键词：
Adopted Affect Alzheimer&apos s Disease Amyloid Amyloid fibers Animal Model Animals Atomic Force Microscopy Atrophic Award Binding Biochemical Biological Assay Brain Cell Culture Techniques Cell Line Cell model Cells Cerebrum Cessation of life Characteristics Corpus striatum structure Cultured Cells Deposition Detection Digestion Disease Disease Progression Disease model Functional disorder Human Huntington Disease In Vitro Individual Infection Inherited Laboratories Laboratory Animal Models Light Mentors Modeling Molecular Molecular Conformation Mus Nerve Degeneration Neurodegenerative Disorders Neurons Otsuka brand of aripiprazole Parkinson Disease Pathogenesis Pathogenicity Pathway interactions Patients Peptide Hydrolases Peptides Phase Play Principal Investigator Prion Diseases Prions Process Property Protein Conformation Protein Fragment Proteins Recombinant Proteins Recombinants Research Resistance Role Sampling Seeds Staging Structure System Techniques Thioflavin T Toes Work amyloid formation base brain tissue conformer disease phenotype human Huntingtin protein human tissue induced pluripotent stem cell monomer mouse model polyglutamine prion-based programs protein aggregate protein misfolding relating to nervous system research study stem

项目摘要

Aberrant folding of proteins is a feature comnnon to many neurodegenerative diseases, including prion disease, Huntington's disease (HD), Alzheimer's disease, and others. Misfolded proteins may adopt several distinct tertiary and quaternary conformations, both in terms of their stable, end states, and in terms of meta-stable intermediate states they acquire as they progress from monomers to large aggregated protein deposits in the brain. Identification of specific protein conformations involved in disease processes is lacking for many neurodegenerative diseases, although we have made progress in this regard for prion disease during the mentored phase ofthis award (Colby et.al., PNAS 2009; Colby etal., PLoS Pathog. 2010). We have begun to elucidate the manner in which biophysical properties of misfolded proteins correlate to and even directly result in specific disease phenotypes. in HD however, the role of misfolded proteins in pathogenesis is controversial and much work remains to be done to establish the conformational basis of this disease. In the independent phase of this award, we will apply the same approaches we have leamed over the past several years studying prion disease to HD in order to identify specific conformations ofthe huntingtin protein that result in neurodegeneration. Using peptides, recombinant protein, and misfolded proteins isolated from laboratory animal models and cell culture models, as well as HD patients, we will assess the extent to which the huntingtin protein is capable of acquiring diverse conformations. We will then determine if any of these conformations are capable of initiating neural death or dysfunction in cell and animal models. In a complementary approach, we will also devielop a technique for detecting misfolded huntingtin based on the propensity of misfolded proteins to nucleate or 'seed' amyloid formation. An analogous assay for detecting prions had femtogram sensitivity (Colby et. al., PNAS, 2007). The ability to sensitively detect misfolded huntingtin will enable us to identify its presence much earlier in disease progression than is currently possible. A better understanding ofthe molecular basis of Huntington's disease and related neurodegenerative disorders shall aid substantially in the search for a cure.

蛋白质的异常折叠是许多神经退行性疾病的共同特征，包括朊病毒病、亨廷顿舞蹈病 (HD)、阿尔茨海默病等。错误折叠的蛋白质可能采用多种不同的三级和四级构象，无论是在稳定、最终状态还是在当它们从单体发展到大的聚集蛋白时，它们获得亚稳定的中间状态大脑中的沉积物。疾病过程中涉及的特定蛋白质构象的鉴定是尽管我们在朊病毒方面取得了进展，但许多神经退行性疾病缺乏该奖项指导阶段的疾病（Colby 等人，PNAS 2009；Colby 等人，PLoS Pathog。 2010）。我们已经开始阐明错误折叠蛋白质的生物物理特性的方式与特定疾病表型相关，甚至直接导致特定疾病表型。然而，在高清中，错误折叠的角色蛋白质在发病机制中的作用存在争议，仍需做大量工作来确定其构象本病的基础。在该奖项的独立阶段，我们将采用与我们相同的方法过去几年研究朊病毒病的过程中学到了 HD 以确定特定的构象导致神经变性的亨廷顿蛋白。使用肽、重组蛋白和从实验室动物模型和细胞培养模型以及 HD 患者中分离出错误折叠的蛋白质，我们将评估亨廷顿蛋白能够获得不同构象的程度。我们然后将确定这些构象中的任何一个是否能够引发细胞中的神经死亡或功能障碍和动物模型。在补充方法中，我们还将开发一种检测错误折叠的技术亨廷顿蛋白基于错误折叠蛋白质成核或“种子”淀粉样蛋白形成的倾向。一个用于检测朊病毒的类似测定法具有飞克级灵敏度（Colby 等人，PNAS，2007）。有能力灵敏地检测错误折叠的亨廷顿蛋白将使我们能够在疾病的早期发现它的存在比目前可能的进展。更好地了解亨廷顿舞蹈症的分子基础疾病和相关的神经退行性疾病将大大有助于寻找治疗方法。