Mechanisms and functional effects of fibrillar aggregation in the lens alpha-crystallins

晶状体α-晶状体蛋白中纤维聚集的机制和功能效应

• 批准号: 10315329
• 负责人: Russell McFarland
• 金额: $ 3.87万
• 依托单位: PORTLAND STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2025-05-12
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10315329
• 关键词: Mechanisms; functional; effects; fibrillar; aggregation

Project Summary Lens transparency is maintained by the abundance of a pair of small heat-shock proteins, αA- and αB-crystallin. Over time, lens proteins (including the α-crystallins) accumulate chemical damage that lead to destabilized states and the nucleation of light-scattering aggregates, which take on a pathogenic state known as cataracts. Understanding how this age-related process of protein aggregation in the lens occurs, or how to prevent/reverse it by therapeutics, has been hindered by our poor understanding of the pathways of α-crystallin aggregation, and the molecular nature (or structures) that define light-scattering opacities in the eye lens. The most common state of protein aggregation is presumably amorphous in nature (i.e., proteinaceous precipitates). However, recent advancement in the detection of amyloids has also supported the hypothesis that one of the other prominent aggregate pathways in the lens may be amyloidogenesis (or the formation of amyloids). In fact, over 5% of the insoluble extracts of cataract lenses have been detected to be amyloid in nature. Furthermore, I have discovered that subtle chemical changes to α-crystallin structure can both promote amyloid formation in the α-crystallins, as well as lead to the formation of a completely novel type of fibrillar aggregation state of αB-crystallin inside living cells, that potentiate formation of light-scattering aggregation in response to other destabilized proteins. This proposal seeks to: 1) characterize a novel and reversible form of fibrillar aggregation in αB-crystallin; and 2) determine how disruption of a conserved motif present in both αA- and αB-crystallin leads to the enhancement of amyloid formation. To accomplish these aims, I will employ a collaborative and multi-disciplinary approach that leverages the enabling technology of single particle CryoEM, together with various biochemical and biophysical methods to characterize these aggregation states of the α-crystallins. Success in these aims are expected to contribute to our understanding of protein misfolding and aggregation in the eye lens – the hallmark of the world's leading cause of vision loss.

项目摘要 透镜的透明性是由一对小的热休克蛋白--αA-和α B-晶状体蛋白--的丰度来维持的。 随着时间的推移，透镜蛋白（包括α-晶体蛋白）积累化学损伤，导致不稳定状态 以及光散射聚集体的成核，其呈现出被称为白内障的致病状态。 了解透镜中与年龄相关的蛋白质聚集过程是如何发生的，或者如何防止/逆转 由于我们对α-晶体蛋白聚集途径的了解不足， 定义眼睛透镜中光散射不透明的分子性质（或结构）。最常见的状态 蛋白质聚集体的性质可能是无定形的（即，蛋白质沉淀物）。但最近的 淀粉样蛋白检测的进展也支持了另一个重要的假设， 透镜中的聚集途径可以是淀粉样蛋白生成（或淀粉样蛋白的形成）。事实上，超过5%的 已经检测到白内障晶状体的不溶性提取物本质上是淀粉样蛋白。另外，我发现 α-晶体蛋白结构的细微化学变化可以促进α-晶体蛋白中淀粉样蛋白的形成， 并导致α B-晶体蛋白在活体内形成一种全新的纤维状聚集状态 细胞，其响应于其他不稳定的蛋白质而增强光散射聚集的形成。这 该提案旨在：1）表征α B-晶状体蛋白中纤维状聚集的一种新的可逆形式;以及2） 确定αA-和α B-晶状体蛋白中存在的保守基序的破坏如何导致增强 淀粉样蛋白的形成为了实现这些目标，我将采用合作和多学科的方法 利用单粒子CryoEM的使能技术，以及各种生物化学和 生物物理方法来表征α-晶体蛋白的这些聚集状态。这些目标的成功是 有望有助于我们理解蛋白质错误折叠和聚集在眼睛透镜-标志 世界上导致视力丧失的主要原因。