Insulin Degrading Enzyme: Physiological Function and its Spatial and Activity Modulation

胰岛素降解酶：生理功能及其空间和活性调节

• 批准号: 9817333
• 负责人: Louis B. Hersh
• 金额: $ 41.18万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9817333
• 关键词: Address; Affect; Affinity; Alzheimer' s Disease; Amyloid beta-Protein; Binding; Binding Sites; Biological Assay; Catabolism; Cell Fractionation; Cell physiology; Cells; Cultured Cells; Development; Diabetes Mellitus; Disease; Endosomes; Enzyme Activation; Enzymes; Genetic study; Goals; Human Genetics; Hydrolysis; Inositol Phosphates; Insulin; Insulinase; Knowledge; Ligands; Link; Lipids; Membrane; Microscopy; Monitor; Peptide Hydrolases; Peptides; Phosphatidylinositols; Physiological; Play; Regulation; Role; Signal Transduction; Site; System; Techniques; Testing; Yeasts; base; endosome membrane; enzyme activity; enzyme substrate; inhibitor/antagonist; innovation; interest; mutant; novel; phosphatidylinositol 3-phosphate; polyanion; receptor internalization; receptor mediated endocytosis

Insulin-degrading enzyme (IDE, insulysin) is a primarily cytosolic peptidase shown to be important in the catabolism of insulin, the amyloid beta peptide, and likely other signaling and intracellular peptides. Its cellular physiology is therefore of considerable interest in the treatment of disorders such as diabetes and Alzheimer's disease. However the site of IDE action on these peptides, which are internalized into or otherwise present in the endosomal system, is yet to be fully understood. We propose to address the question of how this cytosolic enzyme encounters substrate peptides, like insulin, in the endosomal system, exploring a novel mechanism for IDE subcellular localization to this compartment. In particular, we propose that IDE is trafficked to endosomes by binding to membrane anionic lipids, particularly phosphoinositides, through a polyanion-binding site. In the first aim, we will test this hypothesis by using IDE polyanion site mutants, by altering levels of a key phosphoinositide, and by expressing a phosphoinositide binding competitor. We further propose to study, in the second aim, the participation of endosomal IDE in insulin and amyloid(beta(peptide(catabolism. We will manipulate endosomal IDE levels using mutant forms of the enzyme with reduced endosome localization, by decreasing PtdIns(3)P levels, and by increasing endosomal IDE by fusing it with a PtdIns(3)P- targeting domain. In the third aim, we will study the polyanion-dependent activation of IDE and its role in affecting its catabolism of cytosolic peptides. We will use IDE mutants to test the effect of activation on hydrolysis of peptide substrates identified using a ligand trapping technique and peptidomic analyses. We will also test for activation of cellular IDE by inositol phosphates and other potential endogenous activators. We will use our trapping technique to identify other endogenous effectors. These studies will develop a clearer picture of how IDE carries out its physiological functions and greatly benefit efforts to treat IDE related pathophysiological states.(

胰岛素降解酶（IDE，胰岛素）是一种主要是胞质肽酶 在胰岛素，淀粉样β肽肽以及其他信号传导的分解代谢中很重要 细胞内肽。因此，它的细胞生理学对治疗具有很大的兴趣 糖尿病和阿尔茨海默氏病等疾病。但是，IDE对这些行动的站点 内体内或以其他方式存在于内体系统中的肽尚未 完全理解。我们建议解决这种胞质酶遇到的问题 内体系统中的底物肽，例如胰岛素，探索IDE的新机制 该隔室的亚细胞定位。特别是，我们建议IDE被贩运到 通过与膜阴离子脂质（尤其是磷酸肌醇）结合的内体。 聚鸡结合位点。在第一个目标中，我们将使用IDE Polyanion位点检验该假设 突变体，通过改变关键磷酸肌醇的水平，并通过表达磷酸肌醇 约束竞争者。我们进一步建议在第二个目标中研究 胰岛素和淀粉样蛋白的内体IDE（β（肽）（分解代谢。我们将操纵内体 使用酶的突变形式的IDE水平，并通过降低内体定位， 通过与ptdins融合（3）p-来降低PTDINS（3）P水平，并增加内体IDE 定位域。在第三个目标中，我们将研究IDE和 它在影响其胞质肽的分解代谢中的作用。我们将使用IDE突变体测试效果 使用配体捕获技术鉴定的肽底物的水解激活的激活 肽组分析。我们还将测试肌醇磷酸盐和 其他潜在的内源激活剂。我们将使用陷阱技术来识别其他 内源性效应子。这些研究将对IDE如何实现其进行更清晰的了解 生理功能，并极大地有益于治疗IDE相关的病理生理状态的努力（（