Extended Data Fig. 5: Re-analyses of protein adduction by aHNE and aONE in RKO cells.
From: A modification-centric assessment tool for the performance of chemoproteomic probes
Raw data sets for such re-analyses were retrieved from Ref. 40 (Yang, et al., Anal Chem, 2015) and Ref. 9 (Sun, et al., Mol Cell Proteomics, 2017). a, Schematic of the workflow for quantitative chemoproteomic analyses of dynamic aHNE/aONE-derived protein adducts in RKO cells. Cells were first treated with either aHNE or aONE. After treatment, cells were either harvested immediately and used as controls or placed in probe-free medium for another 1 and 4 h recovery period. The probe-labeled proteomes were digested with trypsin and then biotinylated by click chemistry with the light (L, recovery) or heavy (H, control) labeled UV-cleavable azido-biotin, followed by streptavidin enrichment, photorelease, and LC-MS/MS analysis. Identification and quantification were performed using the pFind studio (See Methods for more details). b-c, Venn diagrams revealing that pChem-based identification of previously unknown PDMs substantially expanded the target spectrum of aHNE (b) and aONE (c). Note, cysteines on those N-term ketoamide peptide adducts (∆307.15) are assigned as aHNE-modified sites, since such a PDM is most likely generated through an intramolecular rearrangement from Cys to N-term. d-e, Dynamics of aHNE- and aONE-based PDMs in RKO cells. d, Violin plots of L/H ratios determined from two types of aHNE-derived protein adducts in dynamic adduction analyses. e, Violin plots of L/H ratios determined from four types of aONE-derived protein adducts in dynamic adduction analyses. Dash lines denote median value and dot lines denote the 25th and 75th percentiles.
