Metastable Crystallins: Structure and Stabilization

亚稳态晶体蛋白：结构和稳定性

• 批准号: 8470982
• 负责人: KRISHNA K SHARMA
• 金额: $ 47.96万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8470982
• 关键词: Accounting; Adult; Aging; Alzheimer' s Disease; Blindness; Body part; Cataract; Cataract Extraction; Cell Culture Techniques; Cells; Client; Complex; Crystallins; Data; Degenerative Disorder; Deposition; Development; Diabetes Mellitus; Disease; Eye; Glaucoma; Goals; Human; Huntington Disease; Implant; Intervention; Intraocular lens implant device; Investigation; Knowledge; Lead; Learning; Malignant Neoplasms; Methods; Modification; Molecular Chaperones; Mutation; Names; Nuclear; Organ; Parkinson Disease; Pathogenesis; Peptides; Play; Protein Conformation; Proteins; Reagent; Recovery; Research; Role; Savings; Site; Site-Directed Mutagenesis; Structure; Systems Analysis; Testing; Therapeutic; Time; Tissues; age related; cost; crosslink; functional restoration; innovation; insight; lens; lens protein; lens transparency; light scattering; macromolecule; mutant; novel; peptide A; prevent; protein aggregate; protein aggregation; protein misfolding; protein protein interaction; public health relevance; research study; stem; synthetic peptide; tool

DESCRIPTION (provided by applicant): Many diverse diseases, including cataract, some types of glaucoma, Alzheimer disease, diabetes and cancer to name only a few, are now known to share the common feature of aggregated, misfolded or modified protein deposits. The pathological hallmark in this group of diseases is protein aggregation and deposition in specific cells, tissues or organs. The pathological similarities indicate that common principles that govern protein interactions underlie protein misfolding degenerative diseases. Cataract is a classic example of a protein misfolding and aggregation disease. Cataract, a major cause of blindness in the world, develops as a result of age-related modifications and aggregation of the lens proteins. ?-crystallin accounts for nearly 40% of the adult lens proteins but its structure-function is yet to be fully understood. The chaperone-like activity of ?-crystallin is believed to play a central role in maintaining lens transparency. During aging, lens crystallins undergo truncation and these modifications correlate with lens crystallin aggregations responsible for light scattering. It is well established that the addition of ?-crystallin to aggregating proteins stops a further increase in aggregation and light scattering. We have demonstrated that specific sequences in ?-crystallin subunits suppress aggregation of denaturing proteins. However, the full potential of ?-crystallin-derived peptides (mini-chaperones) is yet to be realized. To enhance our understanding of the role of non-native interactions in cataract formation, we propose the following specific aims: Aim 1. Identify the sequences involved in abnormal interactions of ?A-crystallin subunits that lead to protein aggregation in human cataract-causing ?A-crystallin mutants (?AR49C, ?AF71V, ?AG98R and ?AR116H) using novel deuterated cross linkers, biotynylated reagents and peptide arrays. Aim 2. Determine the mechanism and efficacy of ?A-crystallin-derived mini-chaperone in preventing the aggregation of metastable ?A- crystallins (mutants and truncated crystallins) and restoring chaperone activity. The specific aims will be accomplished using novel cross-linkers and mass spectrometric methods. We will also use site-directed mutagenesis and cell culture expression and analysis systems to confirm the cross linking data. These innovative studies will give us new insights into potential interventions for protein misfolding diseases, not only of the eye but also of other parts of the body.

描述（由申请人提供）：现在已知许多不同的疾病都具有聚集、错误折叠或修饰的蛋白质沉积物的共同特征，其中仅举几例，包括白内障、某些类型的青光眼、阿尔茨海默病、糖尿病和癌症。这类疾病的病理特征是特定细胞、组织或器官中蛋白质的聚集和沉积。病理学上的相似性表明，控制蛋白质相互作用的共同原则是蛋白质错误折叠退行性疾病的基础。白内障是蛋白质错误折叠和聚集疾病的典型例子。白内障是世界上导致失明的主要原因，是由于晶状体蛋白与年龄相关的改变和聚集而产生的。 β-晶状体蛋白占成人晶状体蛋白的近40%，但其结构功能尚未完全了解。 β-晶状体蛋白的分子伴侣样活性被认为在维持晶状体透明度方面发挥着核心作用。在老化过程中，晶状体蛋白会发生截断，这些修饰与负责光散射的晶状体蛋白聚集相关。众所周知，向聚集蛋白中添加β-晶状体蛋白可以阻止聚集和光散射的进一步增加。我们已经证明β-晶状体蛋白亚基中的特定序列抑制变性蛋白的聚集。然而，β-晶状体蛋白衍生肽（微型伴侣）的全部潜力尚未实现。增强 根据我们对非天然相互作用在白内障形成中的作用的理解，我们提出以下具体目标： 目标 1. 识别参与 ?A- 晶状体蛋白亚基异常相互作用的序列，这些亚基导致人类白内障引起的 ?A- 中蛋白质聚集使用新型氘化交联剂、生物酰化试剂的晶状体蛋白突变体（?AR49C、?AF71V、?AG98R 和 ?AR116H）和肽阵列。目标 2. 确定 α-A-晶状体蛋白衍生的微型伴侣在防止亚稳态 α-晶状体蛋白（突变体和截短的晶状体蛋白）聚集和恢复伴侣活性方面的机制和功效。具体目标将使用新型交联剂和质谱方法来实现。我们还将使用定点诱变和细胞培养表达和分析系统来确认交联数据。这些创新研究将为我们提供对蛋白质错误折叠疾病的潜在干预措施的新见解，不仅针对眼睛，而且针对身体其他部位。