Hydroxyl radical production by H2O2 plus Cu,Zn-superoxide dismutase reflects the activity of free copper released from the oxidatively damaged enzyme

K. Sato, T. Akaike, M. Kohno, M. Ando, H. Maeda

Research output: Contribution to journalArticle

142 Citations (Scopus)

Abstract

To elaborate the catalytic activity of Cu2+ of Cu,Zn-superoxide dismutase (SOD) in the generation of hydroxyl radical (·OH) from H2O2, we investigated the mechanism of inactivation of α1-protease inhibitor (α1- PI), mediated by H2O2 and Cu,Zn-SOD. When α1-PI was incubated with 500 units/ml Cu,Zn-SOD and 1.0 mM H2O2, 60% of anti-elastase activity of α1- PI was lost within 90 min. ESR spin trapping using 5,5-dimethyl-1-pyrroline N-oxide showed that free ·OH was indeed generated in the reaction of Cu,Zn- SOD/H2O2; this was substantiated by the almost complete eradication of ·OH by either ethanol or dimethyl sulfoxide accompanied by the generation of carbon-centered radicals. ·OH production and α1-PI inactivation in the H2O2/SOD system became apparent at 30 min or later. Dimethyl sulfoxide and 5,5-dimethyl-1-pyrroline N-oxide protected inactivation of α1-PI significantly in this system, indicating that α1-PI inactivation was mediated by ·OH. SOD activity decreased rapidly during the reaction with H2O2 for the initial 30 min. Time-dependent changes in the ESR signal of SOD showed the destruction of ligands for Cu2+ in SOD by H2O2 within this initial period. Thus we conclude that inactivation of α1-PI is mediated in the H2O2/Cu,Zn-SOD system via the generation of ·OH by free Cu2+ released from oxidatively damaged SOD.

Original languageEnglish
Pages (from-to)25371-25377
Number of pages7
JournalJournal of Biological Chemistry
Volume267
Issue number35
Publication statusPublished - 1992
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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