Figure 4.

Proteomics experiments with biotinylated ABP analogs to identify ABP targets. (A) IC₅₀ determination of 11RK72 and 11RK73 for UCHL1. (B) Schematic representation of pull-down experiment to identify ABP binding proteins. (C) Confirmation of UCHL1 pull-down with biotinylated ABP analogs by Western blot analysis. WB was performed using UCHL1 and actin antibodies. Actin was used as a loading control and incubated together with UCHL1 antibody in the input sample. (D) iBAQ values of enzymes related to the Ub system identified in the pull-down LC–MSMS experiment averaged over three replicates. (E) iBAQ values of the top-12 highest-ranking proteins from the pull-down LC–MSMS experiment averaged over three replicates. *ES1 has been annotated as glutamine amidotransferase-like class 1 domain-containing protein 3A or 3B (GATD3A or GATD3B). Reference intensities (iBAQ values) of corresponding proteins in wild-type HEK293T cells³⁹ are shown in Supporting Information Figure S4.