Protein Radical Formation Resulting from Eosinophil Peroxidase-catalyzed Oxidation of Sulfite.
Eosinophil peroxidase (EPO) is an abundant heme protein in eosinophils that catalyzes the formation of cytotoxic oxidants implicated in asthma, allergic inflammatory disorders, and cancer. It is known that some proteins with peroxidase activity (horseradish peroxidase and prostaglandin hydroperoxidase) can catalyze oxidation of bisulfite (hydrated sulfur dioxide), leading to the formation of sulfur trioxide anion radical ((.)SO(3)(-)). This free radical further reacts with oxygen to form peroxymonosulfate anion radical ((-)O(3)SOO(.)) and the very reactive sulfate anion radical (SO(4)()), which is nearly as strong an oxidant as the hydroxyl radical. However, the ability of EPO to generate reactive sulfur radicals has not yet been reported. Here we demonstrate that eosinophil peroxidase/H(2)O(2) is able to oxidize bisulfite, ultimately forming the sulfate anion radical (SO(4)()), and that these reactive intermediates can oxidize target proteins to protein radicals, thereby initiating protein oxidation. We used immuno-spin trapping and confocal microscopy to study protein oxidation by EPO/H(2)O(2) in the presence of bisulfite in a pure enzymatic system and in human promyelocytic leukemia HL-60 clone 15 cells, maturated to eosinophils. Polyclonal antiserum raised against the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) detected the presence of DMPO covalently attached to the proteins resulting from the DMPO trapping of protein free radicals. We found that sulfite oxidation mediated by EPO/H(2)O(2) induced the formation of radical-derived DMPO spin-trapped human serum albumin and, to a lesser extent, of DMPO-EPO. These studies suggest that EPO-dependent oxidative damage may play a role in tissue injury in bisulfite-exacerbated eosinophilic inflammatory disorders.
Ranguelova
Kalina
K
Laboratory of Pharmacology, NIEHS, National Institutes of Health, Research Triangle Park, NC 27709, USA. RanguelovaK@niehs.nih.gov
Chatterjee
Saurabh
S
Ehrenshaft
Marilyn
M
Ramirez
Dario C
DC
Summers
Fiona A
FA
Kadiiska
Maria B
MB
Mason
Ronald P
RP
D000838:Anions / Q000737:chemistry; D003497:Cyclic N-Oxides / Q000737:chemistry; D047209:Eosinophil Peroxidase / Q000737:chemistry / Q000378:metabolism*; D005609:Free Radicals; D018922:HL-60 Cells; D006801:Humans; D017665:Hydroxyl Radical; D007700:Kinetics; D018613:Microscopy, Confocal / Q000379:methods; D008954:Models, Biological; D018384:Oxidative Stress; D010100:Oxygen / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D019097:Spin Trapping; D013447:Sulfites / Q000737:chemistry*
D000838:Anions; D003497:Cyclic N-Oxides; D005609:Free Radicals; D011506:Proteins; D013447:Sulfites; D017665:Hydroxyl Radical; C017245:5,5-dimethyl-1-pyrroline-1-oxide; D047209:Eosinophil Peroxidase; D010100:Oxygen