AP-MS analysis of human FUBIeS30/FAU WT and non-cleavable G73,4A and G74V mutants

To discover candidate FUBIeS30 proteases, we performed affinity purification from human cell lysates expressing tandem Strep II-hemagglutinin (StHA)-tagged WT or mutant (G73,74A or G74V) FUBIeS30-StHA constructs. Subsequent analysis of the co-purified proteins by mass spectrometry (LC-MS/MS) identified differentially interacting factors. We found that two deubiquitinases, USP10 and USP36, were enriched on the non-cleavable mutants compared to WT and subsequently confirmed that USP36 is involved in FUBIeS30 processing in vivo.