On the role of bicarbonate in peroxidations catalyzed by Cu,Zn superoxide dismutase

The interaction of Cu,ZnSOD with H2O2 generates an oxidant at the active si te that can then cause either the inactivation of this enzyme or the oxidat ion of a variety of exogenous substrates. We show that the rate of inactiva tion, imposed by 10-mM H2O2 at 25 degrees C and pH 7.2, is not influenced b y 10-mM HCO3-; whereas the oxidation of 2,2'-azino-bis-[3-ethylbenzothiazol ine sulfonate] (ABTS(=)) is virtually completely dependent upon HCO3-. The reduction of the active site Cu(II) by H2O2, which precedes inactivation of the enzyme, occurred at the same rate in phosphate buffer with or without bicarbonate added. These results indicate that HCO3- does not play a role i n facilitating the interaction of H2O2 with the active site copper, but the y can be accommodated by the proposal that HCO3- is oxidized to HCO3., whic h then diffuses from that site and causes the oxidation of substrates, such as ABTS(=), that are too large to traverse the solvent access channel to t he Cu(II). (C) 1999 Elsevier Science Inc.