Quantitative reactivity profiling predicts functional cysteines in proteomes

Abstract

Cysteine is the most intrinsically nucleophilic amino acid in proteins, where its reactivity is tuned to perform diverse biochemical functions. The absence of a consensus sequence that defines functional cysteines in proteins has hindered their discovery and characterization. Here, we describe a proteomics method to quantitatively profile the intrinsic reactivity of cysteine residues en masse directly in native biological systems. Hyperreactivity was a rare feature among cysteines and found to specify a wide range of activities, including nucleophilic and reductive catalysis and sites of oxidative modification. Hyperreactive cysteines were identified in several proteins of uncharacterized function, including a residue conserved across eukaryotic phylogeny that we show is required for yeast viability and involved in iron-sulfur protein biogenesis. Finally, we demonstrate that quantitative reactivity profiling can also form the basis for screening and functional assignment of cysteines in computationally designed proteins, where it discriminated catalytically active from inactive cysteine hydrolase designs.

Identification of Cys255 in HIF‐1α as a novel site for development of covalent inhibitors of HIF‐1α/ARNT PasB domain protein–protein interaction

“…36 Moreover, there are recent reports where cysteine alkylation of specific residues in target proteins has significant therapeutic effect in addition to well-known examples of irreversible kinase inhibitors. For example, the modification of Cys179 in IKKb by CDDO (2-cyano-3,12-dioxooleana-1,9,-dien-28-oic acid) was shown to inhibit IKKb kinase activity, which in turn fails to activate NF-jB.…”

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“…In recent decades, many studies have demonstrated that metabolic disorder of biothiols will lead to extreme damage in human and numerous other animals (Townsend et al, 2003;Reddie and Carroll, 2008;Weerapana et al, 2010). Specifically, a low level of biothiols in vivo may cause slowed growth, liver damage, weight loss, and some other adverse influences (Wang et al, 2005;Duan et al, 2008;Yang et al, 2012), while a high level of biothiols is closely related to Parkinson's, Alzheimer's, and cardiovascular diseases (Heafield et al, 1990;Rusin et al, 2004;Lin et al, 2008;Yang et al, 2011).…”

The Expanding Landscape of the Thiol Redox Proteome

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