Detecting Nitrated Proteins by Proteomic Technologies

Nitration is a posttranslational modification of tyrosine residues of proteins mediated by peroxynitrite (ONOO-). It commonly occurs in neurological and pathological disorders, which involve nitric oxide (NO)-mediated oxidative stress. Nitration of tyrosine or tyrosyl groups of a protein modulates protein function and initiates signal transduction pathways, which lead to alternation of cellular metabolism and functions. Because of its apparent significance, there is an increasing urge to identify nitrated proteins as a bridge to expand our understanding of their involvement in different biological processes. This chapter describes strategies that could be used for rapid screening and detection of nitrated proteins, subsequent resolution, and identification of nitrated proteins and peptides using proteomic technologies. These include two-dimensional gel electrophoresis coupled with Western blotting and matrix-assisted laser desorption/ionization time of flight mass spectrometry, as well as liquid chromatography-linked tandem mass spectrometry.