Regulation and Catalysis of Human Insulin Degrading Enzyme

人胰岛素降解酶的调控与催化

• 批准号: 8212894
• 负责人: WEI-JEN TANG
• 金额: $ 27.63万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-15 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8212894
• 关键词: 10q; Acetylation; Address; Adverse effects; Affect; Allosteric Regulation; Alzheimer' s Disease; Amyloid; Biochemical; Biological Assay; Biological Process; Bradykinin; Brain; CCL3 gene; CCL4 gene; Catalysis; Chemicals; Chicago; Chronic Disease; Complex; Coupled; Cysteine; Data; Development; Diabetes Mellitus; Dimerization; Drug Delivery Systems; Endorphins; Energy Transfer; Enzyme Gene; Enzymes; Funding; Future; Genetic; Goals; Human; Human Chromosomes; Inflammation; Institutes; Insulin; Insulin-Like Growth Factor II; Insulinase; Intermediate Filament Proteins; Kinetics; Knock-out; Late Onset Alzheimer Disease; Lead; Location; Metalloproteases; Methods; Molecular; Molecular Conformation; Mus; Natriuretic Peptides; Non-Insulin-Dependent Diabetes Mellitus; Peptides; Phenotype; Phosphorylation; Physiological; Point Mutation; Post-Translational Protein Processing; Property; Proteins; Proteomics; Rattus; Regulation; Reporting; Research; Rodent; Role; Screening procedure; Series; Single Nucleotide Polymorphism; Structure; Therapeutic; Therapeutic Agents; Transforming Growth Factors; Ubiquitin; Universities; Vimentin; Work; Yang; base; beta-Chemokines; chemokine; design; dimer; driving force; enzyme activity; enzyme mechanism; genetic analysis; human disease; hydroxamate; innovation; insight; islet amyloid polypeptide; monomer; nestin protein; non-drug; overexpression; oxidation; peptidomimetics; single molecule; small molecule; success; tool

DESCRIPTION (provided by applicant): Insulin degrading enzyme (IDE) is an evolutionarily conserved, 110 kDa metalloprotease that is involved in the clearance of insulin and amyloid ? (A?). Accumulating genetic evidence in rodents and humans strongly support the role of IDE in the progression of type 2 diabetes mellitus and Alzheimer's disease. Thus, it is vital to understand the functions, catalytic mechanism, and regulation of IDE from a molecular perspective to develop viable IDE-based therapeutic strategies. We have solved the structures of human IDE in complex with insulin, A?, and other functionally relevant substrates such as natriuretic peptides and proinflammatory chemokines, CCL3/CCL4. IDE has two 55 kDa domains, which form an enclosed catalytic chamber to entrap its substrates. Our structural and biochemical analyses reveal how IDE uses an enclosed catalytic chamber to selectively recognize the global features of its substrates. The long-term goal of this research is (1) to delineate the function(s) of IDE and the mechanism(s) of its regulation and (2) to elucidate the role of IDE in human diseases. The objectives of this application are to understand (1) the molecular basis for the open-closed conformational switch and dynamics of IDE during catalysis and (2) the molecular mechanism for the regulation of IDE. In addition, we will develop potent chemical modulators of IDE to be used as potential therapeutic agents and as tools to address the biological functions of this enzyme. The central hypothesis is that the open-closed conformational switch of IDE is the key regulatory step of IDE that is subject to allosteric regulation by dimerization, posttranslational modifications, cellular factors, and chemical modulators. The rationale for the proposed research is that understanding the regulation and functions of IDE and developing small chemical modulators of IDE will ultimately allow us to better design IDE-based therapeutic strategies specific to certain human diseases such as diabetes, Alzheimer's disease, and inflammation. Guided by our preliminary data, we will study the regulation and functions of IDE in three specific aims: in Aim 1, we will use single molecule Forster resonance energy transfer analyses to address the conformational switches and dynamics that occur during catalysis of IDE and to determine how the catalysis of IDE is regulated. Aim 2 is to use two distinct screening methods to develop potent small molecule compounds that can modulate the activity of IDE and use such compounds to address the biological functions of IDE. Aim 3 is to combine structural, biochemical, and mutational studies to address the molecular basis for the regulation of IDE by dimerization, physiologically relevant cellular factors such as intermediate filament proteins, nestin and vimentin, and by posttranslational modifications such as phosphorylation and acetylation. The proposed research is significant because it will generate new insights in the dynamics and regulation of a key enzyme involved in diabetes and Alzheimer's disease and because it will also lead to the discovery of new chemical leads that can potently modulate this enzyme. The proposed research is innovative because it employs biophysical, biochemical, cellular, and medicinal chemical approaches to investigate the regulation and functions of IDE. PUBLIC HEALTH RELEVANCE: The catalytic activity of IDE is controlled by the open-closed conformational switch, which is regulated by the interaction of IDE with itself (dimerization) and with cellular factors as well as through posttranslational modifications. We propose to use smFRET, biochemical, structural, chemical, and cellular approaches to better understand how IDE is regulated and what its biological functions are. The success of our studies will offer new insights in the regulation of IDE and new tools to explore the therapeutic potential of IDE.

描述（申请人提供）：胰岛素降解酶（IDE）是一种进化保守的110 kDa金属蛋白酶，参与胰岛素和淀粉样蛋白的清除。（A？）在啮齿动物和人类中积累的遗传证据强烈支持IDE在2型糖尿病和阿尔茨海默病进展中的作用。因此，从分子水平了解IDE的功能、催化机制和调控机制，对于开发可行的IDE治疗策略至关重要。我们已经解决了人IDE与胰岛素，A？，和其它功能相关的底物，如利钠肽和促炎趋化因子，CCL 3/CCL 4。IDE具有两个55 kDa的结构域，其形成封闭的催化室以捕获其底物。我们的结构和生化分析揭示了IDE如何使用封闭的催化室来选择性地识别其底物的全局特征。本研究的长期目标是（1）阐明IDE的功能及其调节机制，（2）阐明IDE在人类疾病中的作用。本申请的目的是了解（1）催化过程中IDE的开-闭构象转换和动力学的分子基础和（2）IDE调节的分子机制。此外，我们还将开发IDE的有效化学调节剂，用作潜在的治疗剂和解决这种酶的生物学功能的工具。中心假设是IDE的开-闭构象转换是IDE的关键调节步骤，其受到二聚化、翻译后修饰、细胞因子和化学调节剂的变构调节。这项研究的基本原理是，了解IDE的调节和功能，并开发IDE的小化学调节剂，最终将使我们能够更好地设计针对某些人类疾病（如糖尿病、阿尔茨海默病和炎症）的IDE治疗策略。在我们的初步数据的指导下，我们将研究IDE在三个特定目标的调节和功能：在目标1中，我们将使用单分子Forster共振能量转移分析来解决IDE催化过程中发生的构象开关和动力学，并确定IDE的催化是如何调节的。目的2是使用两种不同的筛选方法来开发可以调节IDE活性的有效小分子化合物，并使用此类化合物来解决IDE的生物学功能。目的3是结合联合收割机的结构，生物化学和突变的研究，以解决IDE的二聚化，生理相关的细胞因子，如中间丝蛋白，巢蛋白和波形蛋白，并通过翻译后修饰，如磷酸化和乙酰化的调节的分子基础。这项研究意义重大，因为它将对糖尿病和阿尔茨海默病中一种关键酶的动力学和调节产生新的见解，还因为它还将导致发现新的化学先导物，可以有效地调节这种酶。这项研究具有创新性，因为它采用生物物理，生物化学，细胞和药物化学方法来研究IDE的调节和功能。 公共卫生关系：IDE的催化活性由开-闭构象开关控制，其通过IDE与自身（二聚化）和与细胞因子的相互作用以及通过翻译后修饰来调节。我们建议使用smFRET，生物化学，结构，化学和细胞的方法，以更好地了解IDE是如何调节和它的生物学功能是什么。我们研究的成功将为IDE的监管提供新的见解，并为探索IDE的治疗潜力提供新的工具。