The Chemistry and Biology of Protein Glycation

蛋白质糖化的化学和生物学

• 批准号: 10461916
• 负责人: Rebecca Scheck
• 金额: $ 30.36万
• 依托单位: TUFTS UNIVERSITY MEDFORD
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10461916
• 关键词: Address; Affect; Age related macular degeneration; Aging; Alzheimer' s Disease; Amino Acid Sequence; Apolipoproteins; Binding Sites; Biochemical; Biological; Biology; Cardiovascular Diseases; Cell Cycle Progression; Cells; Chemicals; Chemistry; Collaborations; Computer Models; Crystallins; Data; Development; Diabetes Mellitus; Disease; Environment; Enzymes; Felis catus; Future; Genetic; Goals; Health; Human; Hyperglycemia; Impairment; Inflammatory; Investigation; Knock-out; Lead; Learning; Link; Malignant Neoplasms; Methods; Molecular; Mutation; Outcome; Pathogenesis; Pathway interactions; Peptide Library; Pharmacology; Play; Point Mutation; Post-Translational Protein Processing; Predisposition; Prevention; Proteins; Proteolysis; Publishing; Reaction; Reporter; Role; Series; Signal Transduction; Site; Stress; Structure; System; Testing; Ubiquitin; Ubiquitination; Variant; Work; adduct; age related; combinatorial; enzyme activity; experimental study; glycation; glycosylation; improved; insight; lens; multicatalytic endopeptidase complex; new therapeutic target; novel; prevent; protein degradation; protein structure; sugar; tool

Project Summary Glycation is a non-enzymatic posttranslational modification (PTM) that occurs through the spontaneous reaction of reducing sugars or sugar metabolites with amino and guanidino groups on proteins. Glycation is a hallmark of many diseases, including diabetes, cardiovascular disease, cancer, Alzheimer's disease, and age- related macular degeneration. However, compared to enzymatic PTMs, the biological role of glycation remains poorly understood. This is because traditional biochemical and chemical biology tools, which inhibit or profile specific enzyme activities, are not well-suited for the study of a PTM that transpires without an enzyme. As a result, new methods that can overcome the challenge of non-enzymatic, spontaneous, chemistry in living cells are critically needed. The overall objectives of this project are to develop chemical methods that can precisely control the glycation of ubiquitin, and to use these to study the functional consequence of glycation on ubiquitin- driven protein degradation. Our specific aims are to (1) identify features of protein sequence that promote glycation outcomes (2) identify features of protein structure that influence glycation outcomes, and (3) determine the impact of glycation on ubiquitin-driven proteolysis. Our central premise is that selective glycation is templated by the combination of sequence and structure that surrounds a reactive site. In support of this premise, our preliminary and published data show that primary sequence can govern both overall glycation levels and specific product distributions, while protein structure sculpts the specific glycation products that form. In our first aim, we examine the contributions of sequence using combinatorial peptide libraries to vary primary sequence and examine the effect on glycation type and extent for each of the four glycated sites we have identified in ubiquitin. These experiments will identify sequences that promote varying extents of total glycation and/or bias the formation of specific glycation products. In our second aim, we describe experiments to further define how structure contributes to glycation outcomes using a series of point mutations that disrupt structure or alter the chemical microenvironment within ubiquitin. We also integrate these data with computational models that will be used to predict glycation sites. Finally, we will use this information to define and validate a series of “dialAGE” ubiquitin variants that can predictably modulate glycation outcomes in living cells by altering susceptibility to glycation, and/or by promoting the formation of specific glycation adducts. In our third aim, we use these tools to uncover the mechanism through which glycation prevents protein degradation by the ubiquitin-proteasome system in living cells. Specifically, we will test the hypothesis that the glycation of ubiquitin itself impairs protein degradation. This work will therefore provide long- sought methods that can explore the functional role of glycation, and will reveal new therapeutic targets for a range of diseases. These studies will dramatically improve our understanding of glycation and will have an immediate impact on our appreciation for how this unique PTM influences human health, aging, and disease.

项目总结