Role of deamidation in human beta-crystallin structure

我们研究的总体目标是确定脱酰胺在透镜中的作用。 在上一个资助周期，我们令人信服地证明，脱酰胺导致 可溶性聚集体，不稳定的b-晶状体蛋白，改变的b-亚基之间的相互作用， 并增加了陪伴蛋白B-晶状体蛋白所需的α-晶状体蛋白。这些结果表明 脱酰胺诱导晶体蛋白不溶的机制，并强烈支持 脱酰胺作用直接导致白内障的形成。 基于我们的研究结果，我们假设脱酰胺降低了晶状体蛋白 稳定性导致最终引发白内障形成的聚集。 目标1中的实验将识别透镜中潜在相关的脱酰胺。接下来我们 将确定先前确定的脱酰胺是否会导致聚集或减少 通过使用体内模型，稳定性直接导致白内障形成。实验 目标2将确定脱酰胺引发聚集的机制， 鉴定b-晶状体蛋白内和与a-伴侣蛋白的改变的相互作用。 在b-晶体蛋白中存在许多脱酰胺位点。建议的实验是 创新之处在于，它们将区分功能相关的网站和 是有害的，使用最先进的方法直接测试脱酰胺作用 in vivo.未来的研究将筛选防止晶体蛋白聚集的药物。

The overall goal of our research is to determine the role of deamidation in the lens. During the last grant cycle, we convincingly demonstrated that deamidation led to soluble aggregates, destabilized b-crystallins, altered interactions between b-subunits, and increased the a-crystallin needed to chaperone b-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 b-crystallins and with the a-chaperone. Numerous deamidation sites exist in the b-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.