Low-Toxicity Sulfonium-Based Probes for Cysteine-Specific Profiling in Live Cells

Abstract

Despite being a low-abundance amino acid, cysteine plays an essential role in regulating protein function and serves as a satisfactory target of post-translational modifications and drug developments. To comprehensively assess reactive-cysteine-containing proteins, the development of chemical proteomic probes to label cysteine residues in human cells is an important objective. Cysteine modification using sulfonium-based probes is a novel method to identify reactive cysteine residues in proteins. Herein, we reported a set of "cysteine-reactive sulfonium-based (C-Sul)" probes to label the reactive cysteine sites in cellular proteins. Notably, water-soluble C-Sul probes have a significantly enhanced stability and cellular uptakes, displaying a high specificity toward reactive cysteines and compatibility with quantitative proteomic profiling. In comparison to the conventional iodoacetamide-based probe, C-Sul particularly has no inhibitory effects on cell viability, enabling its application in proteomic profiling of reactive cysteine residues under biorelevant conditions. We propose C-Sul probes as optimal tools of cysteine profiling for further broadly basic research.

Chemical modification of proteins – challenges and trends at the start of the 2020s

“…124 Propargylsulfonium probes have also been used for proteomics profiling of reactive Cys residues in live cells. 125 They can also undergo amino–yne reactions with Lys residues, as mentioned previously for the affinity-guided MDM4 conjugation ( vide supra ). 45…”

“…124 Propargylsulfonium probes have also been used for proteomics profiling of reactive Cys residues in live cells. 125 They can also undergo amino-yne reactions with Lys residues, as mentioned previously for the affinity-guided MDM4 conjugation (vide supra). 45 Strained dibenzocyclooctynes (DBCOs) undergo SPAAC reactions with azides of engineered proteins, but can also react with Cys residues in thiol-yne reactions.…”

Quantitative Chemoproteomic Methods for Reactive Cysteinome Profiling

“…In particular, the labeling profiles of IA and UK were systematically compared and these two probes showed unique coverage of cysteinome, [51] respectively. Recently, other types of cysteinereactive labeling reagents have also been developed including benziodoxolone, [52] dinitroimidazole, [53] sulfonium, [54] fluorobenzene, [55] vinylheteroarenes, [56] heteroaromatic azoline thioethers (HAT) [57] and heteroaromatic sulfones [58] and etc. From an unexpected off-target reaction during metabolic glycan labeling, acetylated unnatural sugars were found to label cysteines quite selectively in cell lysates [59,60] and its target cysteinome profile was distinct from that obtained by iodoacetamide.…”

Cyclopropenone, Cyclopropeniminium Ion, and Cyclopropenethione as Novel Electrophilic Warheads for Potential Target Discovery of Triple-Negative Breast Cancer

“…Over the past decade, chemical proteomics has emerged as a powerful technology for target discovery, and many new targets have been identified by means of this technology and have been used in drug development. − However, a large fraction of the proteome is still inaccessible with current chemical probes, and these proteomic entities remain defined as undruggable targets. − Covalent inhibitors have received considerable attention due to their remarkable pharmaceutical properties, but the reactive warhead of covalent drugs are limited to α,β-unsaturated amides. , Consequently, it is highly desirable to develop novel chemical probes, especially novel electrophilic warheads that can be exploited in proteome profiling experiments . Recently, several novel reactive groups such as heteroaromatic sulfones, sulfonium ions, 2-sulfonylpyridine, sulfur triazole, 2-methylthio pyridiniumoxazoline ion, and bicyclobutane carboxylic amide, which target nucleophilic residues in proteins, such as cysteine, lysine, and tyrosine have been developed, and many of them have been used in the development of new types of covalent inhibitors and in target discovery . We recently developed a 2H-3-phenyl-azirine, terminal ynamide, and a diaryltetrazole for chemoselective modification of carboxylic acid residues in live cells. − To further expand the toolbox of reactive warheads, we developed here a suite of novel electrophiles for proteome profiling and the potential of developing new types of covalent inhibitors.…”

“…10,11 Consequently, it is highly desirable to develop novel chemical probes, especially novel electrophilic warheads that can be exploited in proteome profiling experiments. 12 Recently, several novel reactive groups such as heteroaromatic sulfones, 13 sulfonium ions, 14 2-sulfonylpyridine, 15 sulfur triazole, 16 2-methylthio pyridiniumoxazoline ion, 17 and bicyclobutane carboxylic amide, 18 which target nucleophilic residues in proteins, such as cysteine, lysine, and tyrosine have been developed, and many of them have been used in the development of new types of covalent inhibitors and in target discovery. 19 We recently developed a 2H-3-phenyl-azirine, terminal ynamide, and a diaryltetrazole for chemoselective modification of carboxylic acid residues in live cells.…”