Potentiometric titrations and oxidation-reduction potentials of manganese and copper-zinc superoxide dismutases

Abstract

Bovine erythrocyte superoxide dismutase and 2 Mn-containing superoxide dismutases were reduced by the indirect coulometric titration method with methylviologen as the mediator-titrant. On the basis of the titration data the Mn-containing superoxide dismutases contain 1 g-atom of metal/mol of enzyme (dimer). E0'' [redox potential] is +0.31 V for the enzyme from Escherichia coli which exhibits a complicated pH dependence above neutral pH. The Bacillus stearothermophilus Mn-containing enzyme has an E0'' of +0.26 V and .DELTA.Em[midpoint potential]/pH is 50 mV. Bovine erythrocyte superoxide dismutase exhibits anomalous behavior in the coulometric titration curves, which is indicative of 2 nonequivalent Cu centers in the enzyme. Addition of K3Fe(CN)6 or K2IrCl6 to the enzyme solution, prior to coulometric titration, indicates that these anions bind preferentially to 1 of the Cu centers.