SELENODEIODINASE PROCESSING BY THE PROTEASOME SYSTEM

蛋白酶体系统处理硒代脱碘酶

• 批准号: 6628589
• 负责人: PHILIP REED LARSEN
• 金额: $ 21.06万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-01 至 2004-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6628589
• 关键词: enzyme activity; fasting; gene mutation; genetically modified animals; iodide peroxidase; laboratory mouse; myocardium; proteasome; protein metabolism; protein sequence; thyroxine; triiodothyronine; ubiquitin; yeasts

DESCRIPTION: (Adapted from the applicant's abstract) This is a new proposal to determine the mechanisms regulating the rate-limiting steps in the ubiquitin proteasome system leading to the controlled degradation of the Types 1, 2 and 3 selenodeiodinases. D2 is a critical enzyme regulating the first step in thyroid hormone action, the conversion of the prohormone T4 to the active hormone, T3. This process occurs intracellularly in brain, pituitary and brown fat in which the T3 produced constitutes a major fraction of the nuclear receptor-bound hormone. Furthermore, it appears that, in humans, unlike in adult rodents, D2 may also generate a significant fraction of plasma T3 by virtue of its wide tissue expression in skeletal muscle. It has been known for a number of years that post-translational regulation of D2 by substrate is a major regulatory step in determining the tissue level of the active enzyme. Recent studies have shown that degradation of D2 occurs in proteasomes, that its half-life is quite short (<1 hr), that its degradation is accelerated 2-fold by exposure to substrate and that this process requires interaction of substrate with a D2 enzyme which has either selenocysteine or cysteine in its active center. To define the regulatory steps in this process, the investigator will use several complementary systems. These include transient expression in HEK-293 cells and a temperature-sensitive CHO cell line containing a mutant UB-1 enzyme. A third strategy will be to exploit the fact that D2 can be expressed in yeast with well-documented mutations in the ubiquitin-proteasome degradation pathway and that it is possible to express both wild type and epitope-labeled protein to evaluate changes in activity those in D2 protein levels. Lastly, the investigator will employ cell-free systems which will allow the study of specific components of the ubiquitin proteasome system in the metabolism of D2 and compare results with those for Dl and D3 which have significantly longer half-lives. From a physiological perspective, it is quite conceivable that the accelerated proteolysis of D2 in human skeletal muscle during starvation or illness could explain the rapid onset of the "low T3 syndrome." This is a well-recognized but poorly understood phenomenon which occurs in every human under food restriction or with significant systemic illness.

描述：（根据申请人的摘要改编）这是一个新建议 确定调节泛素限速步骤的机制 蛋白酶体系统导致类型1、2和3的受控降解 Selenodeiodinases。 D2是调节甲状腺第一步的关键酶 激素作用，激素T4转化为活性激素，T3。 这个过程在大脑，垂体和棕色脂肪中细胞内发生 T3产生的构成了核受体结合的主要部分 激素。此外，看来，在人类中，与成年啮齿动物不同，D2 由于其较宽 骨骼肌的组织表达。它已经闻名了很多年 底物对D2的翻译后调节是主要的调节 确定活性酶的组织水平。最近的研究 表明D2的降解发生在蛋白酶体中，其半衰期是相当的 短（<1小时），通过暴露于2倍的降解加速了 底物，并且此过程需要底物与D2的相互作用 酶在其活性中心具有硒代半胱氨酸或半胱氨酸。到 在此过程中定义监管步骤，研究人员将使用几个 互补系统。这些包括在HEK-293细胞中的瞬时表达和 温度敏感的CHO细胞系，其中包含突变的UB-1酶。第三 策略将是利用D2可以在酵母中表达的事实 泛素 - 蛋白酶体降解途径中有据可查的突变和 可以表达野生型和表位标记的蛋白 评估活性变化D2蛋白水平中的活动。最后， 研究人员将采用无细胞系统，该系统将允许研究 D2代谢中泛素蛋白酶体系统的特定成分 并将结果与​​DL和D3的结果进行比较 半衰期。从生理的角度来看，可以想象 在饥饿期间，D2加速D2的蛋白水解或 疾病可以解释“低T3综合征”的快速发作。这是一个 众所周知但知之甚少的现象发生在每个人中 在食品限制或严重的全身性疾病下。