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. 2021 Jan 15;7(3):eabc8590. doi: 10.1126/sciadv.abc8590

Fig. 5. XIAP ubiquitinates proteins and peptides with unmodified N termini.

Fig. 5

(A) Coomassie staining of purified wild-type or inactive (His467Ala) XIAP. (B) IBs of anti-NAA10, anti-NAA15, and anti-vinculin of input lysates used for DDX47 immunoprecipitations (IP) displayed in (C). (C) In vitro ubiquitination assay on DDX47 immunoprecipitates from HeLa cells treated for 48 hours with NatA RNAi or mock. In vitro ubiquitinations were performed as described in Methods using wild-type or mutant XIAP as E3 ligase. IBs of anti-ubiquitin and anti-DDX47 are shown. (D) Unacetylated and acetylated biotinylated peptides of DIABLO, COMMD10, CASP9, PIGS, and DDX47 bound to streptavidin beads were subjected to in vitro ubiquitination assays with wild-type or mutant XIAP. Streptavidin beads were used as additional control. Anti-ubiquitin and anti-streptavidin IBs are presented.