Crystallin-Derived Anti-Chaperones in the Lens

晶状体中的晶状体蛋白衍生的反伴侣分子

• 批准号: 8306862
• 负责人: KRISHNA K SHARMA
• 金额: $ 36.2万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8306862
• 关键词: Accounting; Age; Aging; Binding Sites; Biochemical; Blindness; Cataract; Cavia; Cell Nucleus; Cells; Cleaved cell; Complex; Crystallins; Disease; Generations; Goals; Health; Human; Hydrogen Peroxide; Hyperbaric Oxygen; In Vitro; Lead; Lens Opacities; Mass Spectrum Analysis; Measures; Mediating; Methods; Mind; Modeling; Molecular; Molecular Chaperones; Molecular Weight; Nuclear; Peptide Hydrolases; Peptides; Physical condensation; Precipitation; Predisposition; Property; Proteins; Proteolysis; Reaction; Relative (related person); Role; Sampling; Senile Cataract; Site; Spatial Distribution; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Structure; Systems Analysis; Techniques; Testing; Time; Toxic effect; Water; age related; aged; analytical tool; in vivo; inhibitor/antagonist; lens; lens protein; lens transparency; method development; novel; prevent; protein aggregate; protein aggregation; protein structure; young adult

DESCRIPTION (provided by applicant): Changes in the structure of crystallins, the major lens protein, are thought to account for the loss of lens transparency that occurs with aging. Aged and cataractous human lenses show increased crystallin proteolysis and aggregation as compared to young, non-cataractous lenses. However, the molecular mechanisms for age-related protein aggregation are not fully understood. We hypothesize that protein aggregation in the aging lens (>40 years old) is initiated by the actions of novel proteases, which recognize a specific sequence in crystallins and are responsible for cleavage of crystallin and generation of crystallin fragments with anti-chaperone activity and toxic properties. The crystallin fragments with anti-chaperone activity interact with 1A- and 1B-crystallins at chaperone sites, contributing to the loss of the 1-crystallin chaperone activity known to occur in aged, less transparent lenses and cataractous lenses. Additionally the interactions between crystallin fragments and intact, modified or truncated crystallins lead to age-related protein aggregation and insolubilization of 1-crystallin in the nuclear region of the lens, resulting in loss of transparency and cataract formation. In support of our hypothesis that crystallin-derived anti-chaperone peptides have a key role in the age-related loss of lens transparency, we have demonstrated that in vivo-generated human lens crystallin fragments generate H2O2, display anti-chaperone activity and induce lens protein aggregation and precipitation. Studies will be performed in both human lenses and in the guinea pig model of age-related cataractogenesis. We propose to undertake studies to identify the origin of crystallin fragments and to characterize the proteolytic mechanisms responsible for the cleavage of crystallins and the generation of anti-chaperone peptides. Using state-of-the-art mass spectrometric tools and analytical techniques, we will expand our understanding of the molecular mechanisms involved in crystallin-derived-peptide mediated protein aggregation and cataractogenesis. The proposed studies will accomplish the following Specific Aims: 1) Investigate the low-molecular weight crystallin fragments present in young, adult, aged and cataractous human lenses and in lenses of normal and hyperbaric oxygen (HBO)-treated guinea pigs. 2) Characterize the crystallin-derived anti-chaperone peptides and determine their ability to induce 1- crystallin aggregation in vitro and ex vivo. 3) Investigate whether sequence-specific proteases or non-enzymatic cleavages are responsible for the generation of crystallin-derived anti-chaperone peptides and isolate and characterize the novel protease involved. PUBLIC HEALTH RELEVANCE: Understanding why the lens so commonly develops a cataract may lead to the development of methods for preventing this common cause of vision loss. During aging, for some unknown reason, the lens begins to accumulate peptides (breakdown products) derived from crystallin (peptides), the major protein in the lens. Over time, these peptides become toxic in the lens as they accumulate, leading to loss of lens transparency and cataract.

描述（由申请人提供）：晶体结构的变化，主要晶状体蛋白，被认为是随着衰老而发生的晶状体透明度的损失。与年轻的非伴晶状体相比，老化和白内障的人透镜显示出晶体蛋白的蛋白水解和聚集的增加。但是，与年龄相关蛋白质聚集的分子机制尚不完全了解。我们假设衰老晶状体（> 40岁）中的蛋白质聚集是由新型蛋白酶的作用引发的，新蛋白酶的作用识别结晶蛋白中的特定序列，并负责结晶蛋白的裂解和具有抗链球酮活性和毒性特性的结晶蛋白片段的产生。具有抗伴侣活性的结晶蛋白片段在伴侣部位与1a-和1b-晶状蛋白相互作用，这导致已知在老年，透明透镜和白内障镜片中丧失已知的1-晶状蛋白酶伴侣活性。另外，结晶蛋白片段与完整，修饰或截短的结晶蛋白之间的相互作用导致与年龄相关的蛋白质聚集以及1-晶状体在晶状体核区域中的不脱纤维化，从而导致透明度和白内障形成的丧失。为了支持我们的假设，即结晶蛋白衍生的抗链蛋白肽在与年龄相关的透镜透明度丧失中具有关键作用，我们已经证明，体内生成的人透镜晶体晶体碎片产生了H2O2，会产生H2O2，显示抗链球酮的活性和诱导透镜蛋白蛋白蛋白质的聚集和沉淀。研究将在人类镜片和与年龄相关的白内垂直发生的豚鼠模型中进行。我们建议进行研究，以鉴定结晶蛋白片段的起源，并表征负责结晶蛋白裂解和抗蛋白肽的产生的蛋白水解机制。使用最先进的质谱工具和分析技术，我们将扩展我们对与结晶蛋白衍生的肽介导的蛋白质聚集和白内障发生有关的分子机制的理解。拟议的研究将完成以下特定目的：1）研究年轻，成人，老年和白内障的人类镜片中存在的低分子体重结晶蛋白片段，以及正常和高压氧（HBO）处理的豚鼠的透镜中。 2）表征结晶蛋白衍生的抗链蛋白肽，并确定其在体外和离体中诱导1-结晶蛋白聚集的能力。 3）研究序列特异性蛋白酶或非酶裂解是否是导致结晶蛋白衍生的抗链蛋白肽的产生，并分离并表征涉及的新蛋白酶。 公共卫生相关性：了解为什么如此普遍发展白内障的镜头可能导致开发防止这种常见视力丧失原因的方法。在衰老过程中，由于某些未知原因，晶状体开始积聚源自晶状体中主要蛋白质蛋白质（肽）的肽（分解产物）。随着时间的流逝，这些肽会在晶状体中积聚在晶状体中，从而导致透镜透明度和白内障的丧失。