Bicarbonate enhances the peroxidase activity of Cu,Zn-superoxide dismutase - Role of carbonate anion radical

We examined the effect of bicarbonate on the peroxidase activity of copper-zinc superoxide dismutase (SOD1), using the nitrite anion as a peroxidase probe. Oxidation of nitrite by the enzyme-bound oxidant results in the formation of the nitrogen dioxide radical, which was measured by monitoring 5-nitro-gamma-tocopherol formation. Results indicate that the presence of bicarbonate is not required for the peroxidase activity of SOD1, as monitored by the SOD1/H2O2-mediated nitration of gamma-tocopherol in the presence of nitrite, However, bicarbonate enhanced SOD1/H2O2-dependent oxidation of tocopherols in the presence and absence of nitrite and dramatically enhanced SOD1/H2O2-mediated oxidation of unsaturated lipid in the presence of nitrite, These results, coupled with the finding that bicarbonate protects against inactivation of SOD1 by H2O2, suggest that SOD1/H2O2 oxidizes the bicarbonate anion to the carbonate radical anion. Thus, the amplification of peroxidase activity of SOD1/H2O2 by bicarbonate is attributed to the intermediary role of the diffusible oxidant, the carbonate radical anion. We conclude that, contrary to a previous report (Sankarapandi, S., and Zweier, J. L. (1999) J, Biol, Chem. 274, 1226-1282), bicarbonate is not required for peroxidase activity mediated by SOD1 and H2O2. However, bicarbonate enhanced the peroxidase activity of SOD1 via formation of a putative carbonate radical anion, Biological implications of the carbonate radical anion in free radical biology are discussed.