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Functional Analysis of O-GlcNAc using Synthetic Protein Chemistry

使用合成蛋白质化学对 O-GlcNAc 进行功能分析

基本信息

批准号：
10671580
负责人：
Matthew Robert Pratt
金额：
$ 34.69万
依托单位：
UNIVERSITY OF SOUTHERN CALIFORNIA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-01 至 2025-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10671580
关键词：
Acetylation Affect Amyloid Animals Biochemical Biochemistry Biological Biology Biophysics Carbohydrates Cellular biology Charcot-Marie-Tooth Disease Chemicals Collaborations Cryoelectron Microscopy Data Development Diabetes Mellitus Diffusion Disease Embryonic Development Excision Functional disorder Funding Goals Grant HSPB1 gene Heat shock proteins Kinetics Knowledge Ligation Malignant Neoplasms Mammals Measures Methods Michigan Modification Molecular Molecular Chaperones Monosaccharides Mus Mutagenesis Mutation Nerve Degeneration Neurodegenerative Disorders Neurons Neuropathy O-GlcNAc transferase Parkinson Disease Pathogenicity Pathway interactions Patients Peripheral Nervous System Diseases Phosphorylation Play Point Mutation Polymorph Post-Translational Protein Processing Protein Chemistry Protein Dynamics Proteins Proteomics Publishing Recombinant Proteins Research Role Serine Serine/Threonine Phosphorylation Silicon Dioxide Site Structure Testing Therapeutic Threonine Toxic effect Western Blotting Work alpha synuclein amyloid formation amyloid structure biophysical properties dark matter experimental study glycosylation human disease in vivo monomer mutant prevent programs protein aggregation protein function public health relevance stereochemistry sugar synthetic protein unnatural amino acids

项目摘要

Modified Project Summary/Abstract (The abstract contained no specific references to the in vivo mouse experiments and therefore is unchanged): O-GlcNAc modification is a dynamic protein-modification that is absolutely required for embryonic development in mammals, and is misregulated in diseases, including diabetes, neurodegeneration and cancer. Although approximately 1,000 proteins are modified by O-GlcNAc, the effects of the vast majority of these modifications on protein function are completely unknown. The long-term goal of our research program is to fill in these missing gaps by determining the biochemical consequences of O-GlcNAc on proteins that are key to human disease. To accomplish this goal, we use a combination of carbohydrate and synthetic protein chemistries to build O-GlcNAc modified proteins for subsequent biological experiments. This chemical approach is uniquely enabling, as it is currently the only way to generate homogeneous and site-specifically O-GlcNAc modified proteins. We have been very successful and have used synthetic proteins to determine that O-GlcNAc has a multifaceted role in preventing the amyloid aggregation of proteins in neurodegenerative diseases. Specifically, we have found that O-GlcNAc both directly inhibits the aggregation of amyloid forming proteins and activates the activity of certain small chaperones. In this proposal we will continue to build on these discoveries. In Aim 1, we will determine how O-GlcNAc inhibits the early stages of α-synuclein amyloid formation. In Aim 2, we will test whether O-GlcNAc alters the structure/toxicity relationships of α-synuclein amyloids. In Aim 3, we examine how O-GlcNAc alters the small heat shock protein interactome. Finally, in Aim 4, we will determine if O-GlcNAc can rescue the activity of mutant chaperones that cause Charcot-Marie-Tooth disease. At the conclusion of these independent aims, we will have further unravelled the mechanisms by which O-GlcNAc inhibits protein aggregation and provided critical data to support the ongoing efforts to target O-GlcNAc therapeutically.

项目摘要/摘要（摘要未具体提及小鼠体内实验，因此保持不变）：O-GlcNAc修饰是一种动态的蛋白质修饰，是哺乳动物胚胎发育所绝对需要的，在糖尿病、神经变性和癌症等疾病中被错误调控。虽然大约有1000种蛋白质被O-GlcNAc修饰，但这些修饰对蛋白质功能的影响是完全未知的。我们研究计划的长期目标是通过确定O-GlcNAc对人类疾病关键蛋白质的生化后果来填补这些缺失的空白。为了实现这一目标，我们使用碳水化合物和合成蛋白质化学的结合来构建O-GlcNAc修饰蛋白，用于后续的生物实验。这种化学方法是唯一可行的，因为它是目前唯一的方法来产生均匀和位点特异性的O-GlcNAc修饰蛋白。我们已经非常成功地使用合成蛋白来确定O-GlcNAc在预防神经退行性疾病中蛋白质的淀粉样蛋白聚集方面具有多方面的作用。具体来说，我们发现O-GlcNAc既直接抑制淀粉样蛋白形成蛋白的聚集，又激活某些小伴侣蛋白的活性。在本提案中，我们将继续以这些发现为基础。在Aim 1中，我们将确定O-GlcNAc如何抑制α-突触核蛋白淀粉样蛋白形成的早期阶段。在Aim 2中，我们将测试O-GlcNAc是否改变α-突触核蛋白淀粉样蛋白的结构/毒性关系。在Aim 3中，我们研究了O-GlcNAc如何改变小热休克蛋白相互作用组。最后，在Aim 4中，我们将确定O-GlcNAc是否可以挽救导致腓骨肌萎蔫病的突变伴侣蛋白的活性。在这些独立目标的结论中，我们将进一步揭示O-GlcNAc抑制蛋白质聚集的机制，并提供关键数据来支持正在进行的针对O-GlcNAc治疗的努力。

项目成果

期刊论文数量（22）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Understanding and exploiting the roles of O-GlcNAc in neurodegenerative diseases.

DOI：
10.1016/j.jbc.2023.105411
发表时间：
2023-12
期刊：
JOURNAL OF BIOLOGICAL CHEMISTRY
影响因子：
4.8
作者：
Pratt, Matthew R.;Vocadlo, David J.
通讯作者：
Vocadlo, David J.

Optimization of Chemoenzymatic Mass Tagging by Strain-Promoted Cycloaddition (SPAAC) for the Determination of O-GlcNAc Stoichiometry by Western Blotting.

DOI：
10.1021/acs.biochem.8b00648
发表时间：
2018-10-09
期刊：
Biochemistry
影响因子：
2.9
作者：
Darabedian N;Thompson JW;Chuh KN;Hsieh-Wilson LC;Pratt MR
通讯作者：
Pratt MR

The E3 ligase adapter cereblon targets the C-terminal cyclic imide degron.

DOI：
10.1038/s41586-022-05333-5
发表时间：
2022-10
期刊：
NATURE
影响因子：
64.8
作者：
Ichikawa, Saki;Flaxman, Hope A.;Xu, Wenqing;Vallavoju, Nandini;Lloyd, Hannah C.;Wang, Binyou;Shen, Dacheng;Pratt, Matthew R.;Woo, Christina M.
通讯作者：
Woo, Christina M.

Mechanistic roles for altered O-GlcNAcylation in neurodegenerative disorders.