Calcium-activated protease in cataract formation

白内障形成中的钙激活蛋白酶

• 批准号: 6710673
• 负责人: THOMAS R SHEARER
• 金额: $ 29.97万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-09-30 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6710673
• 关键词: SDS polyacrylamide gel electrophoresis; X ray crystallography; calcium; calpain; cataract; clinical research; confocal scanning microscopy; endopeptidases; enzyme activity; enzyme induction /repression; gene expression; genetically modified animals; human tissue; laboratory mouse; laboratory rat; lens; lens proteins; mass spectrometry; matrix assisted laser desorption ionization; phosphorylation; polymerase chain reaction; protein isoforms; protein structure function; proteolysis; tissue /cell culture; yeast two hybrid system

DESCRIPTION (provided by applicant): This grant is focused on providing a detailed mechanism for the role of calcium-activated proteases (calpains) in lens. The overall hypothesis being tested is that proteolysis by calpains is essential for limited truncation, tighter packing of truncated beta-crystallins, and rearrangement of the cytoskeletal elements occurring during normal lens fiber differentiation and maturation. During cataract formation, over-activation of calpains by massive influx of calcium produces rapid proteolysis and light scattering because the truncated crystallins are no longer properly ordered. Studies during the latest grant cycle produced several unexpected findings relevant to this hypothesis: Activity of lens-specific calpain Lp82, a splice variant of muscle p94, and not m-calpain, was most strongly associated with cataract formation in young rodents. Further, calpain 10 was discovered as a third ubiquitous calpain in human lens, and calpain 10 was markedly increased in diabetic cataracts in rats. Lp82, Lp85 (a low-abundance splice variant in lens), and calpain 10 may truncate specific lens proteins or cleave proteins at unique sites necessary for proper lens maturation or pathologic cataract formation. To test these ideas, three specific aims will be pursued: 1. Determine the biochemical properties and substrates of Lp82. 2. Determine if p94 variants assume the role of rodent Lp82 in human lenses and are responsible for unexplained proteolysis in human lenses. 3. Define the roles of calpain 10 in human lens. These studies will provide a rational basis for the use of protease inhibitors to prevent cataracts.

描述（由申请人提供）：这项资助的重点是提供钙激活蛋白酶（钙蛋白酶）在晶状体中的作用的详细机制。 正在测试的总体假设是，钙蛋白酶的蛋白水解对于正常晶状体纤维分化和成熟过程中发生的有限截断、截断的β-晶状体蛋白的更紧密堆积以及细胞骨架元素的重排至关重要。 在白内障形成过程中，钙大量流入导致钙蛋白酶过度激活，产生快速蛋白水解和光散射，因为截短的晶状体蛋白不再正确排序。 最新资助周期中的研究产生了与这一假设相关的几个意想不到的发现：晶状体特异性钙蛋白酶 Lp82（肌肉 p94 的剪接变体）的活性，而不是 m-钙蛋白酶，与年轻啮齿类动物白内障的形成密切相关。 此外，钙蛋白酶 10 被发现是人类晶状体中第三种普遍存在的钙蛋白酶，并且钙蛋白酶 10 在糖尿病性白内障大鼠中显着增加。 Lp82、Lp85（晶状体中的低丰度剪接变体）和钙蛋白酶 10 可能会截短特定晶状体蛋白或在晶状体正常成熟或病理性白内障形成所需的独特位点切割蛋白质。 为了检验这些想法，将追求三个具体目标： 1. 确定Lp82的生化特性和底物。 2. 确定 p94 变体是否在人类晶状体中承担啮齿动物 Lp82 的角色，并负责人类晶状体中无法解释的蛋白水解。 3. 定义钙蛋白酶10在人类晶状体中的作用。 这些研究将为使用蛋白酶抑制剂预防白内障提供合理依据。