Iron enzyme ribulose-5-phosphate 3-epimerase in Escherichia coli is rapidly damaged by hydrogen peroxide but can be protected by manganese

Jason M. Sobota, James A. Imlay

Research output: Contribution to journalArticle

Abstract

H2O2 is commonly generated in biological habitats by environmental chemistry and by cellular immune responses. H2O 2 penetrates cells, disrupts metabolism, and blocks growth; it therefore is of interest to identify the major cellular molecules that H 2O2 damages and the strategies by which cells protect themselves from it. We used a strain of Escherichia coli that lacks catalases and peroxidases to impose protracted low-grade H2O2 stress. Physiological analysis indicated that the pentose-phosphate pathway, in particular, was poisoned by submicromolar intracellular H2O 2. Assays determined that ribulose-5-phosphate 3-epimerase (Rpe) was specifically inactivated. In vitro studies demonstrated that Rpe employs a ferrous iron atom as a solvent-exposed cofactor and that H2O 2 rapidly oxidizes this metal in a Fenton reaction. The oxidized iron is released immediately, causing a loss of activity. Most Rpe proteins could be reactivated by remetallation; however, a small fraction of proteins were irreversibly damaged by each oxidation cycle, and so repeated cycles of oxidation and remetallation progressively led to permanent inactivation of the entire Rpe pool. Manganese import and iron sequestration are key elements of the H2O2 stress response, and we found that manganese can activate Rpe in vitro in place of iron, converting the enzyme to a form that is unaffected by H2O2. Indeed, the provision of manganese to H2O2-stressed cells protected Rpe and enabled the pentose-phosphate pathway to retain function. These data indicate that mononuclear iron enzymes can be primary targets of H2O2 stress and that cells adapt by shifting from iron- to manganese-centered metabolism.

Original languageEnglish (US)
Pages (from-to)5402-5407
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume108
Issue number13
DOIs
StatePublished - Mar 29 2011

Keywords

  • Hydroxyl radical
  • MntH
  • Oxidative damage
  • OxyR

ASJC Scopus subject areas

  • General

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