Conole Daniel, Mondal Milon, Majmudar Jaimeen D, Tate Edward W
Department of Chemistry, Imperial College London, London, United Kingdom.
Medicine Design, Pfizer Inc., Cambridge, MA, United States.
Front Chem. 2019 Dec 18;7:876. doi: 10.3389/fchem.2019.00876. eCollection 2019.
Deubiquitinating enzymes (DUBs) function to remove or cleave ubiquitin from post-translationally modified protein substrates. There are about 100 known DUBs in the proteome, and their dysregulation has been implicated a number of disease states, but the specific function of many subclass members remains poorly understood. Activity-based probes (ABPs) react covalently with an active site residue to report on specific enzyme activity, and thus represent a powerful method to evaluate cellular and physiological enzyme function and dynamics. Ubiquitin-based ABPs, such as HA-Ub-VME, an epitope-tagged ubiquitin carrying a C-terminal reactive warhead, are the leading tool for "DUBome" activity profiling. However, these probes are generally cell membrane impermeable, limiting their use to isolated enzymes or lysates. Development of cell-permeable ABPs would allow engagement of DUB enzymes directly within the context of an intact live cell or organism, refining our understanding of physiological and pathological function, and greatly enhancing opportunities for translational research, including target engagement, imaging and biomarker discovery. This mini-review discusses recent developments in small molecule activity-based probes that target DUBs in live cells, and the unique applications of cell-permeable DUB activity-based probes vs. their traditional ubiquitin-based counterparts.
去泛素化酶（DUBs）的功能是从翻译后修饰的蛋白质底物上去除或切割泛素。蛋白质组中已知约有100种DUBs，其失调与多种疾病状态有关，但许多亚类成员的具体功能仍知之甚少。基于活性的探针（ABPs）与活性位点残基发生共价反应以报告特定酶活性，因此是评估细胞和生理酶功能及动态的有力方法。基于泛素的ABPs，如HA-Ub-VME，一种带有C端反应性弹头的表位标签泛素，是“DUBome”活性分析的主要工具。然而，这些探针通常不能透过细胞膜，限制了它们仅用于分离的酶或裂解物。可透过细胞的ABPs的开发将使DUB酶能够直接在完整活细胞或生物体的背景下发挥作用，深化我们对生理和病理功能的理解，并极大地增加转化研究的机会，包括靶点验证、成像和生物标志物发现。本综述讨论了针对活细胞中DUBs的小分子基于活性的探针的最新进展，以及可透过细胞的基于DUB活性的探针相对于传统基于泛素的探针的独特应用。
