Role of deamidation in human beta-crystallin structure

描述（由申请人提供）：我们研究的总体目标是确定脱酰胺在晶状体中的作用。在上一个资助周期中，我们令人信服地证明了脱酰胺导致可溶性聚集体，不稳定？-晶体蛋白，改变相互作用？-亚基，增加？-晶体蛋白需要陪伴？-晶体蛋白这些结果提示了脱酰胺诱导晶体蛋白不溶的机制，并有力地支持脱酰胺直接促进白内障的形成。基于我们的发现，我们假设脱酰胺降低了晶状体的稳定性，导致晶状体聚集，最终引发白内障的形成。Aim 1中的实验将确定晶状体中潜在的相关脱酰胺。接下来，我们将通过体内模型确定先前确定的引起聚集或降低稳定性的脱酰胺是否直接导致白内障形成。Aim 2中的实验将确定脱酰胺触发聚集的机制，通过识别改变的相互作用？-结晶蛋白和-伴侣蛋白大量的脱酰胺位点存在于？-crystallins。拟议的实验是创新的，因为它们将区分功能相关的位点和那些有害的位点，使用最先进的方法直接测试体内脱酰胺的作用。未来的研究将筛选阻止晶体蛋白聚集的药物。公共卫生相关性：白内障是全球致盲的主要原因，也是美国政府医疗保健系统最大的支出之一。我们的研究表明，晶状体中主要的年龄相关的改变，脱酰胺，改变晶状体晶体蛋白的结构和功能，并可能诱发体内与白内障相关的聚集。能够防止脱酰胺引起的晶状体结晶蛋白聚集可能预防白内障并有助于预防其他聚集性疾病。

DESCRIPTION (provided by applicant): The overall goal of our research is to determine the role of deamidation in the lens. During the last funding cycle, we convincingly demonstrated that deamidation led to soluble aggregates, destabilized ?-crystallins, altered interactions between ? -subunits, and increased the ?-crystallin needed to chaperone ?-crystallins. These results suggest a mechanism for deamidation-induced insolubilization of crystallins and strongly support that deamidation contributes directly to cataract formation. Based on our findings, we hypothesize that deamidation decreases crystallin stability leading to aggregation that eventually triggers cataract formation. Experiments in Aim 1 will identify potentially relevant deamidations in the lens. Next, we will determine if deamidations previously identified to cause aggregation or decrease stability directly lead to cataract formation by using an in vivo model. Experiments in Aim 2 will determine the mechanism by which deamidation triggers aggregation, by identifying altered interactions within ? -crystallins and with the ?-chaperone. Numerous deamidation sites exist in the ? -crystallins. The proposed experiments are innovative in that they will distinguish between functionally relevant sites and those that are detrimental, using state-of-the art approaches to directly test the role of deamidation in vivo. Future studies will screen for agents that prevent crystallin aggregation. PUBLIC HEALTH RELEVANCE: Cataracts are the leading cause of blindness worldwide and one of the largest expenses to the U.S. government's health care system. Our research suggests that the major age-related modification in the lens, deamidation, alters structure and function of the lens crystallins and may induce aggregation associated with cataracts in vivo. Being able to prevent deamidation-induced aggregation of lens crystallins may prevent cataracts and help to prevent other aggregation diseases.