Novel Probes for Sirtuins: A Chemical Biology Approach

Sirtuins 的新型探针：化学生物学方法

基本信息

批准号：
9303505
负责人：
Martha A. Hass
金额：
$ 48万
依托单位：
ALBANY COLLEGE OF PHARMACY
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：
2017-04-01 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9303505
关键词：
Academia Active Sites Aging-Related Process Alkynes Apoptosis Applications Grants Biogenesis Biological Biological Models Biological Process Biology Biotin Caloric Restriction Cell Cycle Regulation Cell physiology Cells Chemicals Chemistry Complex Conflict (Psychology)Cues DNA Repair Data Deacetylase Detection Disease Disease Progression Drug Industry Ensure Enzymes Epigenetic Process Event Family Fluorescent Dyes Gene Silencing Genetic Transcription Goals Health Benefit Histones Human Image Immunofluorescence Immunologic In Vitro Investigation Label Lead Libraries Link Lysine Malignant Neoplasms Mammalian Cell Mass Spectrum Analysis Measures Mediating Metabolic Metabolic Diseases Metabolism Microscopy Mitochondria Molecular Biology Multienzyme Complexes Niacinamide Nicotinamide adenine dinucleotide O-Acetyl-ADP-Ribose Outcome Pathogenesis Pathologic Peptides Permeability Photoaffinity Labels Physiological Physiological Processes Play Post-Translational Protein Processing Process Property Protein Isoforms Proteins Proteome Proteomics Recombinants Regulation Reporter Reporting Research Role SIRT1 gene Sampling Signal Transduction Sirtuins Site-Directed Mutagenesis Stimulus Techniques Technology Testing Ultraviolet Rays Validation Western Blotting age related base cellular imaging design enzyme activity experimental study functional group imaging study improved inhibitor/antagonist innovation interest novel novel therapeutics protein expression protein protein interaction response scaffold small molecule targeted treatment therapeutic target tool tool development

项目摘要

PROJECT SUMMARY Sirtuins, also called Class III HDACs, consume stoichiometric amounts of nicotinamide adenine dinucleotide (NAD+) to remove acetyl group from lysine residues and to produce nicotinamide and O-acetyl-ADP-ribose. This intriguing class of enzymes has been implicated in regulating various cellular events and has also been suggested to mediate the beneficial effects of calorie restriction (CR). Sirtuins have been intensely pursued by academia and pharmaceutical industry as therapeutic targets. However, controversies on sirtuin biology also peaked during the last few years because of conflicting results from different research groups. This is partly because these enzymes have been discovered recently, and the intricate interaction loops between sirtuins and other proteins make the characterization of them extremely difficult. One of the daunting tasks is to correlate sirtuin activity to disease pathogenesis. Current molecular biology and proteomics techniques report protein abundance rather than active sirtuin content. Innovative chemical tools that can directly probe the functional state of sirtuins are desperately needed. This grant application proposes to take a highly integrative approach to interrogate the functional state of sirtuins in complex biological samples. Our preliminary results demonstrate the feasibility of this strategy. We have obtained a set of powerful chemical probes that are capable of assessing the active content of sirtuins in model systems. In this proposal we plan to synthesize focused libraries of activity-based chemical probes. The scaffolds are designed for enhanced selectivity and labeling efficiency. The probes that score favorably in labeling of recombinant sirtuins will be subjected to the profiling of whole cell lysate. In complex native proteome, the probe should selectively “highlight” the active sirtuin components. Combined with mass spectrometry based proteomics analysis, this strategy should unveil the functional profile of sirtuins under different physiological and pathological conditions. This will provide information on how abnormal enzyme activity will contribute to disease progression. Furthermore, cell permeable probes will also be employed in cellular imaging study. It will enable the simultaneous detection of functional state and localization and empower the direct analysis of sirtuin function in response to cellular and environmental cues.

项目摘要 Sirtuins，也称为III类HDACS，消耗了烟酰胺腺苷二核苷酸的化学计量量（NAD+）从赖氨酸残留物中去除乙酰基，并产生烟酰胺和O-乙酰基-ADP-核糖。这在调节各种细胞事件时，已经暗示着有趣的酶类 Sirtuins已由学术界和制药行业是治疗靶标。但是，由于结果矛盾来自不同的研究小组。这部分是因为最近发现了这些酶， Sirtuins和其他蛋白质之间的复杂相互作用环使它们的表征极为难的。艰巨的任务之一是将Sirtuin活性与疾病发病机理相关联。电流分子生物学和蛋白质组学技术报告了蛋白质抽象，而不是活性Sirtuin含量。创新的迫切需要可以直接探测Sirtuins功能状态的化学工具。该赠款申请提案采用高度集成的方法来询问Sirtuins的功能状态在复杂的生物样品中。我们的初步结果证明了该策略的可行性。我们有获得了一组强大的化学问题，这些问题能够评估模型中Sirtuins的活性含量系统。在此提案中，我们计划综合基于活动的化学问题的集中库。脚手架旨在提高选择性和标签效率。在标签中得分有利的问题重组Sirtuins将经过全细胞裂解物的分析。在复杂的天然蛋白质组中，探针应有选择地“突出”活动的Sirtuin组件。结合基于质谱的蛋白质组学分析，该策略应在不同的生理和病理学下揭示Sirtuins的功能概况状况。这将提供有关异常酶活性将如何导致疾病进展的信息。此外，在细胞成像研究中也将聘请细胞渗透问题。它将启用简单检测功能状态和定位，并赋予对Sirtuin功能的直接分析对细胞和环境线索的反应。