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J Biol Chem, Vol. 275, Issue 19, 14038-14045, May 12, 2000
From the Biophysics Research Institute, Medical College of
Wisconsin, Milwaukee, Wisconsin 53226
The effect of bicarbonate anion
(HCO3
Bicarbonate Enhances the Hydroxylation, Nitration, and
Peroxidation Reactions Catalyzed by Copper, Zinc Superoxide
Dismutase
INTERMEDIACY OF CARBONATE ANION RADICAL*
) on the peroxidase
activity of copper, zinc superoxide dismutase (SOD1) was investigated using three structurally different probes: 5,5'-dimethyl-1-pyrroline N-oxide (DMPO), tyrosine, and
2,2'-azino-bis-[3-ethylbenzothiazoline]-6-sulfonic acid (ABTS).
Results indicate that HCO3 enhanced
SOD/H2O2-dependent (i)
hydroxylation of DMPO to DMPO-OH as measured by electron spin
resonance, (ii) oxidation and nitration of tyrosine to dityrosine,
nitrotyrosine, and nitrodityrosine as measured by high pressure liquid
chromatography, and (iii) oxidation of ABTS to the ABTS cation radical
as measured by UV-visible spectroscopy. Using oxygen-17-labeled water,
it was determined that the oxygen atom present in the DMPO-OH adduct
originated from H2O and not from
H2O2. This result proves that neither free hydroxyl radical nor enzyme-bound hydroxyl radical was involved in the
hydroxylation of DMPO. We postulate that
HCO3 enhances SOD1 peroxidase
activity via formation of a putative carbonate radical anion. This new
and different perspective on HCO3-mediated oxidative reactions of
SOD1 may help us understand the free radical mechanism of SOD1 and
related mutants linked to amyotrophic lateral sclerosis.
*
This work was supported by National Institutes of Health
Grants RR01008 and HL63119 and by a grant from the Amyothophic Lateral Sclerosis Association.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Biophysics Research
Inst., Medical College of Wisconsin, 8701, Watertown Plank Rd., P.O.
Box 26509, Milwaukee, WI 53226. Tel.: 414-456-4035; Fax: 414-456-6512;
E-mail: balarama@mcw.edu.
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