Fig. 1: Workflow and cysteinome coverage from our HT-LFQ chemoproteomic method.
From: Robust proteome profiling of cysteine-reactive fragments using label-free chemoproteomics
a Schematic of our label-free sample preparation method that allows detection of cysteine-containing peptides from lysates or live cells using a hyperreactive IA-DTB probe. b Data acquisition was performed using an Evosep One and Bruker timsTOF Pro 2, followed by identification and quantification of peptides using Spectronaut. Peptides that were bound by a covalent fragment are expected to show a reduced intensity in compound-treated samples, relative to control samples. c Our method allows detection of high numbers of peptides in HEK293T and Jurkat lysates, providing the opportunity to detect liganding events at over 30,000 cysteine residues from over 8000 proteins (n = 16 DMSO control samples from each lysate). d We see high data completeness of peptide detection, with two-thirds of peptides detected in ≥ 75% of samples, allowing more confident detection of binding events. Data shown here is from HEK293T lysate; see Supplementary Fig. 1c for Jurkat data. e Our detection of cysteine residues from an individual cell lysate (HEK293T; orange) represents approximately ~40% coverage of residues that can be considered to be feasibly detectable, based on their location relative to tryptic cleavages sites (green) and the general detectability of proteins by global proteomics methods (purple), as well as the presence of disulfide bonds and post-translational modifications (see Supplementary Fig. 3). Tryptic peptides were classified as being detectable if they were 7–40 residues (not considering missed cleavages). f Distribution of proteins across protein families (top) and target development levels (bottom), as defined by the ‘Illuminating the Druggable Genome’ programme3,7,35. The colour scheme in this figure follows that used in (e). GPCRs: G protein-coupled receptors; TFs: transcription factors. Parts of both (a) and (b) were created in BioRender. Cawood, E. (2025) https://BioRender.com/m32r739. Source data are provided as a Source Data file.
