Chemical biology approaches to probe signaling by protein lipidation

项目摘要 蛋白质脂化是一种动态的翻译后修饰（PTM），影响亚细胞 运输，辅因子结合亲和力和蛋白质的膜定位，这反过来又 影响下游信号级联。特别地，环S-酰化和环S-脱酰 在特定的半胱氨酸残基的蛋白质正在成为循环脂质水平之间的关键环节， 调节基本生物过程，包括参与细胞生长的过程， 新陈代谢和神经健康。然而，对这种PTM的深入研究在技术上证明， 困难的，在很大程度上是由于缺乏选择性，有效的化学抑制剂的酶 催化它的安装和拆除。拟议的研究计划旨在 产生新的化学技术，即小分子探针和抑制剂， 服务阐明参与调节蛋白S-酰化在正常和 病理生理学背景这些目标将通过两种互补的化学物质来实现 和细胞生物学研究领域。一个领域将涉及测量，操纵， 确定S-酰化的“作者”DHHC的目标。为此，泛主动DHHC 将使用新开发和优化的筛选和选择性分析来鉴定抑制剂 结合合理和计算设计的分子的测定，以及 推定抑制剂库。然后将使用经验证的抑制剂和基于蛋白质组学的方法 确定活细胞中DHHC的特异性蛋白质靶点，以更精确地描述其 参与各种疾病状态。第二个研究领域将利用我们最近验证的 化学工具来研究S-酰化的“橡皮擦”的生物功能，APT，特别是 强调这些酶参与细胞氧化还原稳态和代谢 疾病这一多学科研究计划的预期成果是双重的： 一个收集的化学工具和分析的研究这一重要的PTM，并描述其 在正常和疾病状态下的生物学功能和影响。

Project Summary Protein lipidation is a dynamic post-translational modification (PTM) that affects subcellular trafficking, co-factor binding affinity, and membrane localization of proteins, which in turn influence downstream signaling cascades. In particular, cyclic S-acylation and -deacylation of proteins at specific cysteine residues is emerging as a key link between circulating lipid levels and the regulation of essential biological processes, including those involved in cellular growth, metabolism, and neurological health. In-depth study of this PTM, however, has proven technically difficult, in large part due to the paucity of selective, effective chemical inhibitors for the enzymes that catalyze its installation and removal. The proposed research program is designed to generate novel chemical technologies, namely small molecule probes and inhibitors, in the service of illuminating the involvement of regulated protein S-acylation in both normal and pathophysiological contexts. These goals will be realized through two complementary chemical and cellular biology research areas. One area will involve measuring, manipulating, and determining the targets of the “writers” of S-acylation, DHHCs. To do so, pan-active DHHC inhibitors will be identified using newly developed and optimized screening and selectivity profiling assays in combination with rationally and computationally designed molecules, as well as a library of putative inhibitors. Validated inhibitors and proteomics-based methods will then be used to identify the specific protein targets of DHHCs in live cells to more precisely describe their involvement in various disease states. The second research area will utilize our recently validated chemical tools to study the biological function of the “erasers” of S-acylation, APTs, with particular emphasis on the involvement of these enzymes in cellular redox homeostasis and metabolic disease. The expected outcome of this multidisciplinary research program is two-fold: generating a collection of chemical tools and assays for the study of this important PTM, and describing its biological function and influence in normal and disease states.