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Beyond the Active Site: Structure Informed Novel Regulatory Mechanisms and Functional Modulation of O-GlcNAc Transferase

超越活性位点：O-GlcNAc 转移酶的结构知情新型调控机制和功能调节

基本信息

批准号：
10752894
负责人：
Jiaoyang Jiang
金额：
$ 41.16万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2027-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10752894
关键词：
Active Sites Address Affinity Area Binding Binding Proteins Binding Sites Biochemical Biological Biological Assay Biology Catalytic Domain Cells Chemicals Complex Conserved Sequence Coupled Coupling Development Diabetes Mellitus Disease Enzyme Kinetics Enzymes Funding Human In Vitro Insulin Resistance Investigation Knowledge Ligand Binding Domain Link Malignant Neoplasms Mediating Modification Molecular Molecular Conformation Mutation Nutrient O-GlcNAc transferase Outcome Pathologic Processes Pathway interactions Peptides Phage Display Physiological Processes Protein Glycosylation Proteins Proteome Proteomics Regulation Research Resolution Role Scaffolding Protein Site Stress Structure Substrate Interaction System Techniques Time Warburg Effect cancer cell detection of nutrient drug discovery experimental study glycosylation imaging approach innovation insight interest mutant novel protein complex protein protein interaction response sensor structural biology sugar

项目摘要

Abstract The essential human enzyme O-GlcNAc transferase (OGT) catalyzes a unique type of intracellular protein glycosylation called O-GlcNAcylation. In response to nutrient levels and stress, OGT dynamically regulates a variety of physiological and pathological processes including the “Warburg effect” in cancer cells and insulin resistance in diabetes. Previous studies on the OGT active site have made fundamental discoveries on its catalytic mechanism and substrate interactions. However, how OGT regulates protein- and site-specific O- GlcNAcylation remains unclear. This is due to a number of challenges including: 1) OGT glycosylates thousands of proteins without a conserved sequence motif near the O-GlcNAc modification site, 2) a majority of O- GlcNAcylation sites are found on intrinsically disordered regions (IDRs), 3) OGT typically binds proteins with low/moderate affinity, and 4) a lack of OGT-protein complex structures. In our last funding period, we have made strides in these areas through development of a suite of novel chemical probes that allow us to interrogate OGT specific interactions with low/moderate affinity for structural, proteomic, and biochemical characterizations. This proposal aims to make further conceptual and technical breakthroughs toward addressing these longstanding challenges. It is expected that a better understanding of how OGT interacts with other proteins, particularly through the regions beyond the OGT catalytic site, will be essential for understanding OGT’s functional regulation at protein- and site-specific levels, filling major knowledge gaps between decades of biological observations of OGT’s nutrient sensing and other regulatory roles, and will support the need to specifically modulate OGT functions for biomedical applications.

摘要人体必需酶O-GlcNAc转移酶（OGT）催化一种独特类型的细胞内蛋白糖基化称为O-GlcNAc化。为了响应营养水平和压力，OGT动态调节各种生理和病理过程，包括癌细胞和胰岛素中的“瓦尔堡效应抵抗糖尿病。以前对OGT活性中心的研究已经在其结构上取得了基础性的发现。催化机理和底物相互作用。然而，OGT如何调节蛋白质和位点特异性O- GlcNAc化仍不清楚。这是由于许多挑战，包括：1）OGT糖基化数千在O-GlcNAc修饰位点附近没有保守序列基序的蛋白质，2）大部分O- GlcNAc化位点在固有无序区（IDR）上发现，3）OGT通常结合具有以下结构的蛋白质：低/中等亲和力，和4）缺乏OGT-蛋白质复合物结构。在上一个融资期，我们通过开发一套新的化学探针，使我们能够询问OGT，具有低/中等亲和力的特异性相互作用，用于结构、蛋白质组学和生物化学表征。这该提案旨在进一步实现概念和技术突破，以解决这些长期存在的问题挑战希望更好地了解OGT如何与其他蛋白质相互作用，通过OGT催化位点以外的区域，对于理解OGT的功能调节是必不可少的在蛋白质和位点特异性水平上，填补了几十年来生物学观察之间的主要知识空白， OGT的营养传感和其他调节作用，并将支持需要专门调节OGT 生物医学应用的功能。