Assay of metabolic superoxide production in Escherichia coli

J. A. Imlay, I. Fridovich

Research output: Contribution to journalArticlepeer-review


Superoxide production has been measured in subcellular fractions of SOD-deficient Escherichia coli provided with physiological reductants. Although cytosolic enzyme(s) do generate O-2, the larger portion is produced by autoxidation of components of the respiratory electron-transport chain. At 37°C and with pO2, NADH, and NAD+ levels matching those in vivo, respiring membrane vesicles generate 3 O-2/10,000 electrons transferred. This corresponds to intracellular O-2 production, in glucose-fed cells, of 5 μM/s. The high SOD content of normal cells restricts O-2 accumulation to 2·10-10 M, with a moderate gradient from the membrane to the center of the cell. SOD-deficient mutants achieve a much higher steady-state content of O-2. Rates of superoxide-mediated inactivation of certain enzymes are sufficiently rapid that even 10-10 M O-2 imposes a significant oxidative stress.

Original languageEnglish (US)
Pages (from-to)6957-6965
Number of pages9
JournalJournal of Biological Chemistry
Issue number11
StatePublished - 1991
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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