Biophysical basis for enzyme mediated deglycation in protein repair

蛋白质修复中酶介导的去糖化的生物物理学基础

• 批准号: 10798655
• 负责人: Jennifer Binning
• 金额: $ 10.94万
• 依托单位: H. LEE MOFFITT CANCER CTR & RES INST
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10798655
• 关键词: Acetylation; Address; Affect; Aging; Alzheimer' s Disease; Amines; Atherosclerosis; Bacteria; Binding; Biological; Biological Assay; Biological Markers; Biological Process; Biomedical Research; Biophysics; Carbon; Catalysis; Cell Line; Cells; Chemicals; Chemistry; Complex; Coupling; Databases; Degenerative polyarthritis; Development; Diabetes Mellitus; Diagnosis; Disease; Elements; Enzymatic Biochemistry; Enzyme Kinetics; Enzymes; Event; Excision; Family; Fructosamine; Funding; Gene Expression Regulation; Glucose-6-Phosphate; Goals; Homologous Gene; Human; Hybrids; KRP protein; Kinetics; Knock-out; Life; Ligands; Link; Lysine; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Mediating; Methylation; Modification; Molecular; Natural regeneration; Non-Insulin-Dependent Diabetes Mellitus; Organism; Pathway interactions; Pentosephosphate Pathway; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Physiological; Polysaccharides; Post-Translational Protein Processing; Process; Protein Tyrosine Phosphatase; Proteins; Proteome; Reaction; Regulation; Resolution; Ribose; Risk; Role; Site; Structure; Substrate Specificity; Systems Biology; Techniques; Trees; Ubiquitination; Visualization; Work; Writing; biophysical properties; experimental study; glycation; human disease; improved; in vivo; inorganic phosphate; insight; interdisciplinary approach; knock-down; link protein; nervous system disorder; prevent; protein degradation; protein protein interaction; protein purification; protein structure; repaired; structural biology; sugar

Project Summary/Abstract Organisms across all domains of life decorate their protein molecules with an incredible diversity of chemical modifications. Modifications on proteins are critical for their function, affecting protein structure, stability, and interaction partners. Many of the proteins and the enzymes that read, write, and erase these modifications are closely tied to human diseases ranging from neurological disorders to cancer to type 2 diabetes. While these proteins and pathways can be targets to treat these diseases, we lack a high-resolution, mechanistic understanding of how the cell installs, recognizes, and leverages certain post-translational modifications, specifically ubiquitination and spontaneous, non-enzymatic modifications. Our lab is working to understand how protein-protein interactions dynamically regulate post-translational modifications to alter proteome landscape and impact human disease. Protein glycation is an understudied post-translational modification that arises when a sugar covalently attaches to a primary amine. This process occurs spontaneously under normal physiological conditions and is a bio-marker in aging and the development, or worsening, of diseases such as diabetes, Alzheimer's disease, osteoarthritis, and atherosclerosis. Early glycation events are reversible and represent one of the few protein repair mechanisms in the cell. Deglycation is mediated by an unusual “hybrid” kinase/deglycase called Fructosamine-3-kinase (FN3K). FN3K facilitates the removal of protein-linked glycans by directly phosphorylating the attached sugar and destabilizing the sugar-protein linkage. FN3K and FN3K homologs are found in all branches of the tree of life. The glycation of intracellular proteins is not well studied, yet the conservation of FN3K and FN3K-related proteins underscores an important biological role for these enzymes. In this project, my lab will use a multidisciplinary approach, including techniques and expertise in structural biology, enzymology, and systems biology, to address sharply focused mechanistic questions regarding FN3K-mediate protein repair. We hypothesize that an improved mechanistic understanding of FN3K will reveal new biological insight into this ancient repair process, and that we can leverage this insight to better diagnose and treat diseases associated with elevated glycation. In order to distinguish our contributions from those of others, we will integrate reductionist and global approaches to develop a deeper and more complete understanding of the regulation and repair of glycated proteins. Over the five-year funding period, the goals of this project are to: (i) determine the structural and biophysical basis for FN3K-mediated protein repair (ii) systematically characterize the binding kinetics and enzymatic activity of FN3K and FN3K-RP on diverse substrates; (iii) identify sites-specific FN3K deglycation sites and their potential cross-talk with other PTMs. The successful completion of this work will establish the molecular mechanisms that govern the protein deglycation repair process and will ultimately provide needed breakthroughs in biomedical research.

项目摘要/摘要 生命所有领域的生物都以令人难以置信的多样性来装饰其蛋白质分子 化学修饰。对蛋白质的修饰对于其功能至关重要，影响蛋白质结构，稳定性， 和互动伙伴。读，写和擦除这些修饰的许多蛋白质和酶 与从神经疾病到癌症再到2型糖尿病的人类疾病紧密相关。而这些 蛋白质和途径可以是治疗这些疾病的靶标，我们缺乏高分辨率的机械 了解细胞如何安装，识别和利用某些翻译后修改， 特别是泛素化和赞助商，非酶修饰。我们的实验室正在努力了解如何 蛋白质 - 蛋白质相互作用动态调节翻译后修饰以改变蛋白质组景观 并影响人类疾病。 蛋白质糖化是一种经过理解的翻译后修饰，当糖共价时会产生 附着在原胺上。此过程是在正常生理条件下自发发生的，是 糖尿病，阿尔茨海默氏病等疾病的衰老与发育或令人担忧的生物标志物， 骨关节炎和动脉粥样硬化。早期糖基化事件是可逆的，代表了少数蛋白质之一 细胞中的修复机制。脱糖化是由称为的不寻常的“混合”激酶/脱胶酶介导的 果糖胺-3-激酶（FN3K）。 FN3K促进直接通过直接去除蛋白质连接的聚糖 磷酸化附着的糖并破坏糖蛋白键的稳定。 FN3K和FN3K同源物是 在生命之树的所有分支中发现。细胞内蛋白的糖基化糖基糖素的研究不好，但是 FN3K和FN3K相关蛋白的保护强调了这些酶的重要生物学作用。在 这个项目，我的实验室将使用多学科方法，包括结构生物学方面的技术和专业知识， 酶学和系统生物学，以解决有关FN3K-媒体的强烈集中的机械问题 蛋白质修复。我们假设对FN3K的机械理解的改进将揭示新的生物学 洞悉这一古老的维修过程，我们可以利用这种洞察力来更好地诊断和治疗疾病 与糖基化升高有关。为了区分我们的贡献与他人的贡献，我们将整合 还原主义者和全球方法，以对监管和 修复糖化蛋白。在五年的资金期内，该项目的目标是：（i）确定 FN3K介导的蛋白质修复（II）的结构和生物物理基础有系统地表征结合 FN3K和FN3K-RP对潜水员底物的动力学和酶活性； （iii）识别特定地点的FN3K 退化位点及其与其他PTM的潜在串扰。这项工作的成功完成将 建立控制蛋白质度修复过程的分子机制，并最终提供 生物医学研究所需的突破。