PADI4-Cit - Precision bifunctional peptide tools to understand protein citrullination

瓜氨酸蛋白是一种翻译后修饰（PTM），参与表观遗传基因调控和先天免疫应答等多种生物学功能。它的失调与多种疾病的进展有关。尽管瓜氨酸具有这些重要作用，但我们对瓜氨酸的认识仍处于起步阶段。哺乳动物体内的瓜氨酸酶由肽基精氨酸脱亚胺酶（PADI）家族的五种混杂同工酶催化，它们表现出重叠的底物特异性和多种组织表达模式。这种复杂的地形使得精确了解瓜氨酸的影响具有挑战性，目前还没有在单一底物水平上探索瓜氨酸的一般策略。为了剖析这个复杂的系统，我的目标是开发双功能化学工具，将PADI酶招募到特定的底物和基因上，从而能够以目前不可能的精确度和特异性水平研究瓜氨酸对蛋白质功能和基因表达的影响。我将集中在PADI 4，主要的核成员的PADI，生产方法，以精确地了解生化作用的单一瓜氨酸事件在细胞为基础的背景下，在底物水平。我的新双功能工具将用于解决两个相关的目标：了解瓜氨酸对1）单个蛋白质底物的影响，以及2）基因调控。这项工作的结果应该阐明这种相对知之甚少的PTM，为瓜氨酸对蛋白质功能的影响提供新的见解，并为未来研究其他底物和PADI的应用提供蓝图，有可能为治疗干预提供新的途径。

Protein citrullination is a posttranslational modification (PTM) involved in a wide range of biological functions including epigeneticgene regulation and the innate immune response. Its dysregulation is implicated in the progress of diverse pathogeneses. Despitethese important roles, our understanding of citrullination remains in its infancy. Citrullination in mammals is catalysed by fivepromiscuous isozymes from the peptidylarginine deiminase (PADI) family, which display overlapping substrate specificities and arange of tissue expression patterns. This complex landscape makes precise understanding of the effects of citrullination challenging,with no general strategies currently available to explore citrullination at the single substrate level. To dissect this complicated system,I aim to develop bifunctional chemical tools to recruit PADI enzymes to specific substrates and genes, allowing investigation of theeffects of citrullination on protein function and gene expression with a level of precision and specificity not currently possible. I willfocus on PADI4, the primarily nuclear member of the PADIs, producing methods to precisely understand the biochemical effects ofsingle citrullination events in a cell-based context at the substrate level. My new bifunctional tools will be used to address two relatedgoals: understanding the effect of citrullination on 1) individual protein substrates, and 2) gene regulation. Results from this workshould shed light on this relatively poorly understood PTM, providing new insight into the effects of citrullination on protein functionand a blueprint for future applications to study other substrates and PADIs, with the potential to provide new avenues for therapeuticintervention.