Proteomic analysis of oxidative modifications in human atherosclerotic plaques and control arteries

Cardiovascular disease is a major cause of worldwide mortality. A prime underlying cause is atherosclerotic plaque formation at specific arterial locations. Considerable evidence links plaques with chronic inflammation and leukocyte activation, with this potentially resulting in enhanced oxidant-mediated damage to arterial proteins. In this study, human carotid artery plaques and control tissue (superior thyroid artery) were analyzed by liquid chromatography-mass spectrometry (LC-MS/MS) for the presence of post-translational modifications (PTMs) generated by redox reactions on proteins. Of 7150 proteins detected across all samples, 2724 carried at least one PTM, with these including mono- and di-oxidation at methionine, tryptophan and histidine, conversion of cysteine to cysteic acid, chlorination and nitration of tyrosine and tryptophan, as well as carbamylation of lysine and N-terminal amines. Higher PTM levels were detected on proteins from plaques compared to control artery tissue, but similar modification types were detected. Elevated concentrations of PTMs were detected in plaques classified as unstable compared to stable, consistent with PTMs being associated with plaque instability. The levels of several PTMs correlate with the concentration of the neutrophil-derived heme protein myeloperoxidase, implicating this enzyme and its oxidants in protein damage, though direct causality has not been established except for chlorination. Large numbers of PTMs were detected on key metabolic proteins including those involved in glycolysis/gluconeogenesis, the pentose phosphate pathway, the tricarboxylic acid (TCA) cycle and oxidative phosphorylation. Particularly high levels were detected on glycolytic enzymes, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and Complex V subunits consistent with perturbations to cellular metabolism in atherosclerotic plaques.