Developing Mass Spectrometry-based Approaches to Characterize Mono- and Poly(ADP-ribosyl)ated Proteomes

项目总结 翻译后修饰(PTM)涉及将化学基团添加到蛋白质上， 调节修饰蛋白质的结构或功能以协调细胞过程。 ADP-核糖化，即将二磷酸腺苷核糖加到蛋白质上，存在于 单体(聚合)和聚合(聚合)形式，允许更复杂的调节。 ADP-核糖化形式的合成和降解之间的平衡是 调节细胞过程。这种平衡的破坏会导致人类疾病，如癌症和 神经退行性变。然而，关于各自的贡献仍有许多需要了解的地方 MAR化和PAR化对生理过程和疾病的调节 发病机制。基于质谱学(MS)的蛋白质组学方法是 调查健康和疾病状态下的PTM。先前发表的蛋白质组学方法 研究ADP-核糖化已成功地确定了氨基酸的修饰位点 具有不同的化学性质。然而，所有这些方法都不能鉴定ADP-核糖化 与每个站点关联的表单。本提案旨在开发一种基于MS的蛋白质组学方法 在蛋白质组水平上鉴定ADP-核糖化的位置和形式。保存信息 关于修饰部位的ADP-核糖化形式，我的方法将：(A)保持ADP- 在浓缩过程中完整的核糖化作用或(B)确定氨基酸是MAR化的还是 双音节。有了一个工具来区分每个部位的ADP-核糖化形式，我们可以 确定健康和健康人群中MAR化和PAR化蛋白的潜在功能差异 疾病状态。最终，这个工作流程将提供对生物功能的有价值的见解 生理和疾病状态下ADP-核糖化的不同形式。

PROJECT SUMMARY Post-translational modifications (PTMs) involve the addition of chemical groups to proteins, modulating the structures or functions of the modified proteins to coordinate cellular processes. ADP-ribosylation, the addition of adenosine diphosphate ribose to proteins, exists in both a monomeric (MARylation) and polymeric (PARylation) form, allowing for more complex regulation. The balance between the synthesis and degradation of ADP-ribosylation forms is fundamental for regulating cellular processes. Disruption of this balance leads to human diseases like cancer and neurodegeneration. However, much remains to be learned about the respective contributions of MARylation and PARylation to the regulation of physiological processes and disease pathogenesis. Mass spectrometry (MS)-based proteomics methods are the go-to techniques for investigating PTMs in healthy and disease states. Previously published proteomics methods studying ADP-ribosylation have been successful in identifying modification sites on amino acids of varied chemical properties. However, all of these methods fail to identify the ADP-ribosylation form associated with each site. This proposal aims to develop a MS-based proteomics method to identify the site and form of ADP-ribosylation on a proteome-wide level. To preserve information about the form of ADP-ribosylation at modification sites, my method will either (a) keep ADP- ribosylation intact during enrichment or (b) determine whether the amino acid is MARylated or PARylated. With a tool to distinguish between the ADP-ribosylation forms at each site, we can identify potential functional differences of MARylated and PARylated proteins in healthy and disease states. Ultimately, this workflow will provide valuable insights into the biological functions of the different forms of ADP-ribosylation in physiological and disease states.