Ubiquitylation, a biochemical modification made to proteins, is involved in multiple cellular processes, including protein degradation, and its dysregulation plays a role in several cancers. The enzymes that add ubiquitin groups to proteins, known as ubiquitin ligases, are an emerging class of drug targets. Even though 600 E3 ubiquitin ligases have been identified in humans, their substrates remain largely unknown and existing methods for identifying these targets have limitations. Researchers led by Ludwig Harvard’s Hai-Tsang Huang and William Sellers reported in a March issue of Nature Chemical Biology their development of a method, named E3-substrate tagging by ubiquitin biotinylation (E-STUB), that broadly and accurately identifies targets of E3 ubiquitin ligases. Their method specifically labels ubiquitylated substrates in proximity to an E3 ligase of interest with biotin to enable their subsequent identification. The researchers applied E-STUB to characterize the actions of protein degraders (i.e., PROTACs and molecular glues), revealing direct as well as collateral targets—including ubiquitylation events that do not lead to substrate degradation. These results suggest E-STUB could aid new strategies for drug design, like inducing ubiquitylation of proteins central to disease processes that aren’t amenable to binding by inhibitory drugs. Beyond that, E-STUB has great potential as a tool for the functional study of E3 ligases, advancing our understanding of their roles in human health and disease.
Ubiquitin-specific proximity labeling for the identification of E3 ligase substrates
Nature Chemical Biology, 2024 March 21