Protein cross-linking during oxidative denaturation of methaemoglobin

Incubation of methaemoglobin in vitro for up to 4 h led to prominent cross-linking of globin subunits. This process was inhibited by superoxide dismutase, catalase and metal-chelator like diethylenetriamine penta-acetic acid but not by scavengers of hydroxyl radicals. Methaemoglobin in solution produced superoxide radical. It is suggested that damage to globin subunits is mediated by active oxygen species like hydroxyl radical involving a 'site-specific mechanism'. The formation of microprecipitates in incubated samples of methaemoglobin was also inhibited by anti-oxidant enzymes and transition metal-chelator indicating again the involvement of oxygen free-radicals. Such oxidative denaturation of methaemoglobin progresses independent of any formation of hemichrome and may have important physiological significance.