Quantitative reactivity profiling predicts functional cysteines in proteomes. Nature

The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Nature (Impact Factor: 41.46). 11/2010; 468(7325):790-5. DOI: 10.1038/nature09472
Source: PubMed


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 profile quantitatively the intrinsic reactivity of cysteine residues en masse directly in native biological systems. Hyper-reactivity was a rare feature among cysteines and it was found to specify a wide range of activities, including nucleophilic and reductive catalysis and sites of oxidative modification. Hyper-reactive 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 is involved in iron-sulphur protein biogenesis. We also demonstrate that quantitative reactivity profiling can form the basis for screening and functional assignment of cysteines in computationally designed proteins, where it discriminated catalytically active from inactive cysteine hydrolase designs.

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Available from: Sagar D Khare
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    • "Cysteine (Cys), an amino acid in living organisms plays a crucial role in numerous biological systems. Cys is a component of proteins, a source of sulfide in iron–sulfur clusters, and a part of the antioxidant glutathione (GSH) [1] [2]. Abnormal levels of Cys have been linked to several serious diseases, such as edema, hair depigmentation, liver damage, slowed growth and skin lesions [3]. "
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    • "The DUB inhibitory activity is likely to be due to the electrophilic nature of these compounds coupled with the fact that the vast majority of DUBs are cysteine proteases that are sensitive to electrophilic attack. Generally, cysteine residues in proteins are nucleophilic and tend to show high levels of variation with regard to their reactivity to electrophiles (Codreanu et al., 2014; Weerapana et al., 2010). ␣,␤-unsaturated ketones are generally considered as relatively soft electrophiles and are speculated to display selective reactivity towards a specific subset of cysteine thiolates in proteins (Aldini et al., 2006). "
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    • "Redox-Modified Cysteine Residues Occur Outside Common Functional Domains To further explore possible underlying biochemical and biophysical factors that promote specificity of post-translational cysteine modifications, an in-depth structural and biochemical analysis of the proteomes was performed. We did not identify a specific primary sequence motif associated with the cysteine modifications , consistent with previous analyses of large datasets including those of reactive cysteine residues (defined by the reactivity toward iodoacetamide) (Marino and Gladyshev, 2010b; Weerapana et al., 2010). However, the frequency of certain amino acids within a 13-amino-acid window surrounding the modified cysteine residues revealed that cysteine, glutamine, serine, and tryptophan are generally under-represented while alanine, histidine, isoleucine, lysine, methionine, and valine are generally over-represented amino acids (Figure S1). "
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