A powerful new method called E3-substrate tagging by ubiquitin biotinylation (E-STUB) has been developed for identifying the direct ubiquitylated targets of E3 ubiquitin ligases.

Developed by Huang et al., E-STUB employs a ubiquitin variant with a biotin acceptor and an E3 fused to a biotin ligase, enabling biotinylation of proximal ubiquitylated substrates. E-STUB revealed the relationship between ubiquitylation and degradation for multikinase degraders and demonstrated collateral ubiquitylation of corepressor complexes. It discovered physiological substrates of CRBN and VHL using ligase mutants.

The Aurora Ultimate 25×75 C18 UHPLC column enabled sensitive detection and high-resolution separation of peptides for in-depth mass spectrometric analysis.

Publication
Nature Chemical Biology

Authors

Hai-Tsang Huang, Ryan J. Lumpkin, Ryan W. Tsai, Shuyao Su, Xu Zhao, Yuan Xiong, James Chen, Nada Mageed, Katherine A. Donovan, Eric S. Fischer, & William R. Sellers

Title

Ubiquitin-specific proximity labeling for the identification of E3 ligase substrates