The Identification of Protein S-Nitrosocysteine

Abstract

Nitric oxide (NO•) bioactivity regulates cellular function in most major mammalian organ systems. NO signaling either through soluble guanylate cyclase activation and cGMP production, or S-nitrosylation, the modification of cysteine residues to form S-nitrosocysteine, are currently the two most well studied nitric oxide signaling pathways. The emergence of S-nitrosylation as an intricate participant in cellular signaling is evidenced by the identification of endogenously S-nitrosylated proteins, including the NMDA receptor (1), cas-pase-3 (2), GAPDH (3), and NF-κB (4), which all participate in signal transduction pathways that regulate the fine balance between cell survival and cell death. The identification of novel S-nitrosylated proteins has traditionally stemmed from the observation that nitric oxide could modulate a protein’s function, e.g. its enzymatic activity, and that these effects were often recapitulated when thiol reactive compounds such N-ethylmaleimide (NEM) or organomercury derivatives such as p-choloromercuribenzoic acid (PCMB) were used. Moreover, if the observed effects of nitric oxide could be reversed by reducantants such as dithiothreitol (DTT), cysteine modification was suspected. Researchers would then undertake the task of mutational analysis to identify which critical cysteine residue(s) was responsible for the observed effects.