Structural profiling of endogenous S-nitrosocysteine residues reveals unique features that accommodate diverse mechanisms for protein S-nitrosylation

Proc Natl Acad Sci U S A. 2010 Sep 28;107(39):16958-63. doi: 10.1073/pnas.1008036107. Epub 2010 Sep 13.


S-nitrosylation, the selective posttranslational modification of protein cysteine residues to form S-nitrosocysteine, is one of the molecular mechanisms by which nitric oxide influences diverse biological functions. In this study, unique MS-based proteomic approaches precisely pinpointed the site of S-nitrosylation in 328 peptides in 192 proteins endogenously modified in WT mouse liver. Structural analyses revealed that S-nitrosylated cysteine residues were equally distributed in hydrophobic and hydrophilic areas of proteins with an average predicted pK(a) of 10.01 ± 2.1. S-nitrosylation sites were over-represented in α-helices and under-represented in coils as compared with unmodified cysteine residues in the same proteins (χ(2) test, P < 0.02). A quantile-quantile probability plot indicated that the distribution of S-nitrosocysteine residues was skewed toward larger surface accessible areas compared with the unmodified cysteine residues in the same proteins. Seventy percent of the S-nitrosylated cysteine residues were surrounded by negatively or positively charged amino acids within a 6-Å distance. The location of cysteine residues in α-helices and coils in highly accessible surfaces bordered by charged amino acids implies site directed S-nitrosylation mediated by protein-protein or small molecule interactions. Moreover, 13 modified cysteine residues were coordinated with metals and 15 metalloproteins were endogenously modified supporting metal-catalyzed S-nitrosylation mechanisms. Collectively, the endogenous S-nitrosoproteome in the liver has structural features that accommodate multiple mechanisms for selective site-directed S-nitrosylation.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Cysteine / analogs & derivatives*
  • Cysteine / analysis
  • Cysteine / metabolism
  • Hydrophobic and Hydrophilic Interactions
  • Liver / metabolism*
  • Mice
  • Molecular Sequence Data
  • Protein Processing, Post-Translational*
  • Proteins / chemistry
  • Proteins / metabolism*
  • Proteome
  • Proteomics
  • S-Nitrosothiols / analysis
  • S-Nitrosothiols / metabolism*


  • Proteins
  • Proteome
  • S-Nitrosothiols
  • S-nitrosocysteine
  • Cysteine