Extended Data Fig. 3: iDeg-1 causes selective IDO1 depletion by polyubiquitination of lysine 389 of IDO1.
From: Monovalent pseudo-natural products supercharge degradation of IDO1 by its native E3 KLHDC3
a,b, Influence of iDeg-1 on the protein levels of DOCK-8 and RHOBTB3 in HEK293T cells treated for 6 h or 24 h. Representative immunoblots (one of n = 3 biological replicates) (a) and quantification of band intensities from a (b). Mean values ± SD, n = 3 biological replicates. VCL: vinculin. c-e, IFN-γ-stimulated BxPC3 cells were treated with iDeg-1 or DMSO as a control for 6 h prior to lysate preparation. IDO1 was immunoprecipitated and the enriched fraction was digested and analysed by MS. c, IDO1 protein sequence. Amino acids in bold illustrate regions identified in the MS experiment. K389: the identified ubiquitination site of IDO1. Due to incomplete sequence coverage in the MS experiment, ubiquitination of additional lysines cannot be excluded. d, Measured relative peptide intensity for IDO1 and polyubiquitin-C. Mean values ± SD, n = 3 biological replicates. e, Ubiquitin protein sequence. Identified peptides are displayed in bold. The identified ubiquitination site K48 is indicated. K6, K11, K29 and K33 in ubiquitin could not be identified due to incomplete sequence coverage in the MS experiment.
