ABSTRACT
The "peroxidase" activity of the copper-zinc superoxide dismutase is a poorly sustained activity because of the competing inactivation of the enzyme. New evidence suggests that the bound oxidant may be partitioning between oxidizing the enzyme or oxidizing small anions. At constant peroxide, nitrite and azide only partially protect the enzyme (50%) against loss of copper(I) and inactivation up to one anion per copper. Beyond that level, there is no further protection. Bicarbonate ion also protects, but larger amounts are required. These data suggest that there is significant oxidation of the enzyme even in the presence of the small anions and therefore the formation of the bound oxidant cannot be sustained in a true catalytic process.
Subject(s)
Anions , Hydrogen Peroxide/metabolism , Superoxide Dismutase/metabolism , Animals , Binding Sites , Carbon/chemistry , Carbonates/chemistry , Cattle , Cyclic N-Oxides/chemistry , Free Radicals , Hydrogen Peroxide/chemistry , Hydrogen-Ion Concentration , Liver/enzymology , Models, Chemical , Spectrophotometry , Superoxide Dismutase/chemistry , Time FactorsABSTRACT
This work summarizes observations from numerous investigators on the reaction of the copper-zinc superoxide dismutase with hydrogen peroxide at physiological pH in order to propose a likely sequence of events that leads to 2-oxo-histidine formation, copper loss, inactivation, and random and site-specific peptide fragmentation. New data is presented for the bovine liver enzyme that indicate copper is lost as the copper(I) form which immediately reacts with bathocuproine disulfonate to form the characteristic complex that absorbs at 485 nm. Studies in TRIS buffer ruled out the loss of copper(II) followed by reduction of the high potential copper(II)-bathocuproine disulfonate complex by buffer because TRIS is known not to reduce this complex. The rate of loss of copper(I) is not affected by the spin trap, 5,5'-dimethylpyrolline-N-oxide (DMPO), nor by replacing oxygen with argon in the reaction. In addition, changes in the native electrophoretic pattern that are correlated with copper loss and not peptide fragmentation are also unaffected by DMPO, argon, EDTA, or DTPA. These data are taken as indirect evidence that the formation of 2-oxo-histidine is the first oxidative event, unaffected by DMPO, that occurs at the bound oxidant and leads to loss of copper(I). Peptide fragmentation and the peroxidative activity of the dismutase are discussed in light of these observations.
