Catalase-Like Antioxidant Activity is Unaltered in Hypochlorous Acid Oxidized Horse Heart Myoglobin
Gulfam Ahmad,
Belal Chami,
Mary El Kazzi,
Xiaosuo Wang,
Maria Tereza S. Moreira,
Natasha Hamilton,
Aung Min Maw,
Thomas W. Hambly,
Paul K. Witting
Affiliations
Gulfam Ahmad
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Belal Chami
Discipline of Oral Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Mary El Kazzi
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Xiaosuo Wang
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Maria Tereza S. Moreira
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Natasha Hamilton
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Aung Min Maw
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Thomas W. Hambly
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Paul K. Witting
Redox Biology Group, Discipline of Pathology, Faculty of Medicine and Health, Charles Perkins Centre, The University of Sydney, Sydney, NSW 2006, Australia
Activated neutrophils release myeloperoxidase that produces the potent oxidant hypochlorous acid (HOCl). Exposure of the oxygen transport protein horse heart myoglobin (hhMb) to HOCl inhibits Iron III (Fe(III))-heme reduction by cytochrome b5 to oxygen-binding Iron II (Fe(II))Mb. Pathological concentrations of HOCl yielded myoglobin oxidation products of increased electrophoretic mobility and markedly different UV/Vis absorbance. Mass analysis indicated HOCl caused successive mass increases of 16 a.m.u., consistent serial addition of molecular oxygen to the protein. By contrast, parallel analysis of protein chlorination by quantitative mass spectrometry revealed a comparatively minor increase in the 3-chlorotyrosine/tyrosine ratio. Pre-treatment of hhMb with HOCl affected the peroxidase reaction between the hemoprotein and H2O2 as judged by a HOCl dose-dependent decrease in spin-trapped tyrosyl radical detected by electron paramagnetic resonance (EPR) spectroscopy and the rate constant of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS) oxidation. By contrast, Mb catalase-like antioxidant activity remained unchanged under the same conditions. Notably, HOCl-modification of Mb decreased the rate of ferric-to-ferrous Mb reduction by a cytochrome b5 reductase system. Taken together, these data indicate oxidizing HOCl promotes Mb oxidation but not chlorination and that oxidized Mb shows altered Mb peroxidase-like activity and diminished rates of one-electron reduction by cytochrome b5 reductase, possibly affecting oxygen storage and transport however, Mb-catalase-like antioxidant activity remains unchanged.
