Mechanisms of Protein Aging in Normal and Cataractous Lenses

摘要 白内障是世界范围内致盲的主要原因。白内障晶状体摘除术 对本港经济造成沉重的财政负担，亦有副作用。此外，委员会认为， 预计到2050年，美国白内障的患病率将增加一倍以上。 虽然正在开发新的治疗方法来预防或逆转早期白内障， 对白内障发生的潜在机制的理解仍然不清楚。很长的- 我们研究的长期目标是确定老化过程中透镜蛋白的修饰， 白内障的发生，以确定蛋白质老化机制和发展的方法， 延迟或逆转混浊。最近，我们发现了多种年龄依赖性，非- 导致与年龄相关的透镜蛋白质修饰和 白内障相关的透镜蛋白交联。所涉及的透镜蛋白质和氨基酸具有 已经确定;然而，机械细节仍然没有解决。此外，我们还观察到， 在老化的透镜组织中，可溶性透镜蛋白质显著地重新分布到膜部分。 同样，这种蛋白质转变的操作机制仍然未知。我们的假设是 特异性透镜蛋白-蛋白交联、肽/脂质改变和晶状体蛋白与透镜的结合 膜是引起白内障的。为了验证这个假设我们将使用最先进的成像技术 质谱和蛋白质组学/脂质组学方法，以进一步确定年龄相关的， 白内障特异性修饰，并定义发生化学反应的条件， 确定化学机理。具体而言，我们建议：1）定义蛋白质位点， 白内障人晶状体中蛋白质交联的化学，2）定义白内障特异性 人类晶状体中的肽和脂质变化，以及3）阐明导致 随着年龄增长和白内障形成，透镜蛋白膜结合增加。拟议 这些实验有望为蛋白质老化提供新的机制细节， 为新的白内障治疗方法的开发提供信息，同时也指导治疗方法的开发， 其他衰老和蛋白质聚集疾病。

ABSTRACT Cataract is the leading cause of blindness worldwide. Surgical removal of cataractous lenses places an enormous financial burden on our economy and is not without side effects. Moreover, the prevalence of cataract in the U.S. is expected to more than double by the year 2050. Although new therapies are being developed to prevent or reverse early cataract, our understanding of the underlying mechanisms of cataractogenesis remains unclear. The long- term goals of our research are to identify modifications to lens proteins during aging and cataractogenesis in order to define protein aging mechanisms and to develop ways to prevent, delay, or reverse opacification. Recently, we have identified multiple age-dependent, non- enzymatic biochemical processes that result in both age-related lens protein modification and cataract-related lens protein crosslinking. The lens proteins and amino acids involved have been identified; however, mechanistic details remain unresolved. In addition, we have observed a dramatic re-distribution of soluble lens proteins to the membrane fraction in aged lens tissue. Again, the operative mechanism(s) of this protein shift remains unknown. Our hypothesis is that specific lens protein-protein crosslinks, peptide/lipid alterations, and crystallin binding to lens membranes are cataractogenic. To test this hypothesis we will employ state-of-the-art imaging mass spectrometry and proteomics/lipidomics methodology to further define age-related and cataract-specific modifications and define conditions upon which the chemistry occurs to determine chemical mechanism. Specifically we propose to: 1) define the protein sites and chemistry of protein crosslinks in cataractous human lenses and, 2) define cataract-specific peptide and lipid changes in human lenses, and 3) elucidate the molecular events that lead to increased lens protein membrane binding with age and cataract formation. The proposed experiments are expected to provide new mechanistic details on protein aging that will not only inform the development of new cataract treatments, but also guide therapeutic development for other aging and protein aggregation diseases.