Bicarbonate enhances the hydroxylation, nitration, and peroxidation reactions catalyzed by copper, zinc superoxide dismutase - Intermediacy of carbonate anion radical

The effect of bicarbonate anion (HCO3-) on the peroxidase activity of coppe r, zinc superoxide dismutase (SOD1) was investigated using three structural ly different probes: 5,5'-dimethyl-1-pyrroline N-oxide (DMPO), tyrosine, an d 2,2'-azino-bis- [3-ethylbenzothiazolinel-6-sulfonic acid (ABTS), Results indicate that HCO3- enhanced SOD/H2O2-dependent (i) hydroxylation of DMPO t o DMPO-OH as measured by electron spin resonance, (ii) oxidation and nitrat ion of tyrosine to dityrosine, nitrotyrosine, and nitrodityrosine as measur ed by high pressure liquid chromatography, and (iii) oxidation of ARTS to t he ARTS cation radical as measured by UV-visible spectroscopy. Using oxygen -17-labeled water, it was determined that the oxygen atom present in the DM PO-OH adduct originated from H2O and not from H2O2. This result proves that neither free hydroxyl radical nor enzyme-bound hydroxyl radical was involv ed in the hydroxylation of DMPO. We postulate that HCO3- enhances SOD1 pero xidase activity via formation of a putative carbonate radical anion. This n ew and different perspective on HCO3--mediated oxidative reactions of SOD1 may help us understand the free radical mechanism of SOD1 and related mutan ts linked to amyotrophic lateral sclerosis.