Radical acetylation of 2'-deoxyguanosine and L-histidine coupled to the reaction of diacetyl with peroxynitrite in aerated medium.
Diacetyl, like other alpha-dicarbonyl compounds, is reportedly cytotoxic and genotoxic. A food and cigarette contaminant, it is related with alcohol hepatotoxicity and lung disease. Peroxynitrite is a potent oxidant formed in vivo by the diffusion-controlled reaction of the superoxide radical anion with nitric oxide, which is able to form adducts with carbon dioxide and carbonyl compounds. Here, we investigate the nucleophilic addition of peroxynitrite to diacetyl forming acetyl radicals, whose reaction with molecular oxygen leads to acetate. Peroxynitrite is shown to react with diacetyl in phosphate buffer (bell-shaped pH profile with maximum at 7.2) at a very high rate constant ( k 2 = 1.0 x 10 (4) M (-1) s (-1)) when compared with monocarbonyl substrates ( k 2 < 10 (3) M (-1) s (-1)). Phosphate ions (100-500 mM) do not affect the rate of spontaneous peroxynitrite decay, but the H 2PO 4 (-) anion catalyzes the nucleophilic addition of the peroxynitrite anion to diacetyl. The intermediacy of acetyl radicals is suggested by a three-line spectrum ( a N = a H = 0.83 mT) obtained by EPR spin trapping of the reaction mixture with 2-methyl-2-nitrosopropane. The peroxynitrite reaction is accompanied by concentration-dependent oxygen uptake. Stoichiometric amounts of acetate from millimolar amounts of peroxynitrite and diacetyl were obtained under nonlimiting conditions of dissolved oxygen. In the presence of either l-histidine or 2'-deoxyguanosine, the peroxynitrite/diacetyl system afforded the corresponding acetylated molecules identified by HPLC-MS ( n ). These studies provide evidence that the peroxynitrite/diacetyl reaction yields acetyl radicals and raise the hypothesis that protein and DNA nonenzymatic acetylation may occur in cells and be implicated in aging and metabolic disorders in which oxygen and nitrogen reactive species are putatively involved.
Massari
Júlio
J
Departamento de Bioquímica and Departamento de Química Fundamental, Instituto de Química, Universidade de São Paulo, C.P. 26077, 05513-970 São Paulo, SP, Brazil.
Fujiy
Débora E
DE
Dutra
Fernando
F
Vaz
Sandra M
SM
Costa
Ana C O
AC
Micke
Gustavo A
GA
Tavares
Marina F M
MF
Tokikawa
Rita
R
Assunção
Nilson A
NA
Bechara
Etelvino J H
EJ
D000107:Acetylation; D003849:Deoxyguanosine / Q000737:chemistry*; D003931:Diacetyl / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry*; D010100:Oxygen / Q000737:chemistry; D030421:Peroxynitrous Acid / Q000737:chemistry*
D030421:Peroxynitrous Acid; D006639:Histidine; D003849:Deoxyguanosine; D003931:Diacetyl; D010100:Oxygen