title: 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 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.
authors.affiliation: The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
authors.forename: Eranthie
authors.initials: E
authors.lastname: Weerapana
authors.forename: Chu
authors.initials: C
authors.lastname: Wang
authors.forename: Gabriel M
authors.initials: GM
authors.lastname: Simon
authors.forename: Florian
authors.initials: F
authors.lastname: Richter
authors.forename: Sagar
authors.initials: S
authors.lastname: Khare
authors.forename: Myles B D
authors.initials: MB
authors.lastname: Dillon
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Bachovchin
authors.forename: Kerri
authors.initials: K
authors.lastname: Mowen
authors.forename: David
authors.initials: D
authors.lastname: Baker
authors.forename: Benjamin F
authors.initials: BF
authors.lastname: Cravatt
mesh_terms: D000818:Animals; D055162:Biocatalysis; D045744:Cell Line, Tumor; D017124:Conserved Sequence; D003545:Cysteine / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D006867:Hydrolases / Q000737:chemistry / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000096:biosynthesis; D008099:Liver / Q000378:metabolism; D051379:Mice; D009206:Myocardium / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D015202:Protein Engineering; D011492:Protein Hydrolysates; D011484:Protein-Arginine N-Methyltransferases / Q000737:chemistry / Q000378:metabolism; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D020543:Proteome / Q000737:chemistry* / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: C502064:FAM9B protein, human; D007506:Iron-Sulfur Proteins; D009687:Nuclear Proteins; D011492:Protein Hydrolysates; D011506:Proteins; D020543:Proteome; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; C555077:YHR122W protein, S cerevisiae; C007978:hydrolysin; C099988:PRMT1 protein, human; D011484:Protein-Arginine N-Methyltransferases; D006867:Hydrolases; D003545:Cysteine