Kinetics of the oxidation of reduced Cu,Zn-superoxide dismutase by peroxymonocarbonate

Kinetic evidence is reported for the role of the peroxymonocarbonate, HOOCO2−, as an oxidant for reduced Cu,Zn-superoxide dismutase-Cu(I) (SOD1) during the peroxidase activity of the enzyme. The formation of this reactive oxygen species results from the equilibrium between hydrogen peroxide and bicarbonate. Recently, peroxymonocarbonate has been proposed to be a key substrate for reduced SOD1 and has been shown to oxidize SOD1-Cu(I) to SOD1-Cu(II) much faster than H2O2. We have reinvestigated the kinetics of the reaction between SOD1-Cu(I) and HOOCO2− by using conventional stopped-flow spectrophotometry and obtained a second-order rate constant of k=1600±100 M−1 s−1 for SOD1-Cu(I) oxidation by HOOCO2−. Our results demonstrate that peroxymonocarbonate oxidizes SOD1-Cu(I) to SOD1-Cu(II) and is in turn reduced to the carbonate anion radical. It is proposed that the dissociation of His61 from the active site Cu(I) in SOD-Cu(I) contributes to this chemistry by facilitating the binding of larger anions, such as peroxymonocarbonate.

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► Direct optical measurement of SOD1-Cu(I) oxidation by HOOCO2−.
► HOOCO2− formed from CO2 and H2O2 is the precursor to the carbonate anion radical.
► Other mechanisms proposed for SOD1 peroxidase activity are examined.