Yeast ubiquitin ligase LC-MS/MS

Ubiquitin ligases (E3s) serves as a key regulator for ubiquitylation-mediated pathway. The identification of the corresponding relationship between E3 and its substrates is challenging but required for understanding the regulatory network of ubiquitylation. The low abundance of ubiquitinated conjugates and high redundancy of E3s-substrates regulation made screening pretty hard. Herein, we combined SILAC-based quantitative proteomics with two contrary genetic methods (overexpression and knockout) in theory for E3 (Hrt3, F-box subunit of SCF complex) substrates screening.