Inactivation and nitration of human superoxide dismutase (SOD) by fluxes of nitric oxide and superoxide.
Human recombinant MnSOD and CuZnSOD were both inactivated when exposed to simultaneous fluxes of superoxide (JO(2)(*-)) and nitric oxide (J*NO). The inactivation was also observed with varying J*NO/JO(2)(*-) ratios. Protein-derived radicals were detected in both CuZn and MnSOD by immuno-spin trapping. The formation of protein radicals was followed by tyrosine nitration in the case of MnSOD. When MnSOD was exposed to J*NO and JO(2)(*-) in the presence of uric acid, a scavenger of peroxynitrite-derived free radicals, nitration was decreased but inactivation was not prevented. On the other hand, glutathione, known to react with both peroxynitrite and nitrogen dioxide, totally protected MnSOD from inactivation and nitration on addition of authentic peroxynitrite but, notably, it was only partially inhibitory in the presence of the more biologically relevant J*NO and JO(2)(*-). The data are consistent with the direct reaction of peroxynitrite with the Mn center and a metal-catalyzed nitration of Tyr-34 in MnSOD. In this context, we propose that inactivation is also occurring through a *NO-dependent nitration mechanism. Our results help to rationalize MnSOD tyrosine nitration observed in inflammatory conditions in vivo in the presence of low molecular weight scavengers such as glutathione that otherwise would completely consume nitrogen dioxide and prevent nitration reactions.
Demicheli
Verónica
V
Departamento de Bioquímica, Facultad de Medicina, Universidad de la República, Avda. Gral. Flores 2125, 11800 Montevideo, Uruguay.
Quijano
Celia
C
Alvarez
Beatriz
B
Radi
Rafael
R
D003198:Computer Simulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D009569:Nitric Oxide / Q000378:metabolism*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013096:Spermine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013482:Superoxide Dismutase / Q000037:antagonists & inhibitors*; D013481:Superoxides / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D013481:Superoxides; C091861:spermine nitric oxide complex; D013096:Spermine; D009569:Nitric Oxide; D013482:Superoxide Dismutase