Roles of respiratory oxidases in protecting Escherichia coli K12 from oxidative stress (original) (raw)

Abstract

Isogenic strains of Escherichia coli that were defective in either of the two major aerobic terminal respiratory oxidases (cytochromes bo′ and bd) or in the putative third oxidase (cytochrome _bd_-II) were studied to elucidate role(s) for oxidases in protecting cells from oxidative stress in the form of H2O2 and paraquat. Exponential phase cultures of all three oxidase mutants exhibited a greater decline in cell viability when exposed to H2O2 stress compared to the isogenic parent wild-type strain. Cytochrome bo′ mutants showed the greatest sensitivity to H2O2 under all conditions studied indicating that this oxidase was crucial for protection from H2O2 in E. coli. Cell killing of all oxidase mutants by H2O2 was by an uncharacterized mechanism (mode 2 killing) with cell growth rate affected. The expression of Φ(katG-lacZ), an indicator of intracellular H2O2, was 2-fold higher in a cydAB::kan mutant compared to the wild-type strain at low H2O2 concentrations (< 100 μM) suggesting that cytochrome bd mutants were experiencing higher intracellular levels of H2O2. Protein fusions to the three oxidase genes demonstrated that expression of genes encoding cytochrome bd, but not cytochrome bo′ or cytochrome _bd_-II was increased in the presence of external H2O2. This increase in expression of Φ(cydA-lacZ) by H2O2 was further enhanced in a _cyo::_kan mutant. The level of cytochrome bd determined spectrally and Φ(cydA-lacZ) expression was 5-fold and 2-fold higher respectively in an rpoS mutant compared to isogenic wild-type cells suggesting that RpoS was a negative regulator of cytochrome bd. Whether the effect of RpoS is direct or indirect remains to be determined.

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Author notes

1. Jorge Membrillo-Hernández
   Present address: Departamento de Biología Molecular Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Mexico City, Mexico

Authors and Affiliations

1. Department of Microbiology, Otago School of Medical Sciences, University of Otago, P.O. Box 56, Dunedin, New Zealand
   Arne Lindqvist
2. Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts, 02115, USA
   Jorge Membrillo-Hernández
3. Department of Molecular Biology and Biotechnology, The University of Sheffield, Sheffield, S10 2TN, UK
   Robert K. Poole
4. Department of Microbiology, Otago School of Medical Sciences, University of Otago, P.O. Box 56, Dunedin, New Zealand
   Gregory M. Cook

Authors

1. Arne Lindqvist
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2. Jorge Membrillo-Hernández
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3. Robert K. Poole
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4. Gregory M. Cook
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Correspondence toGregory M. Cook.

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Lindqvist, A., Membrillo-Hernández, J., Poole, R.K. et al. Roles of respiratory oxidases in protecting Escherichia coli K12 from oxidative stress.Antonie Van Leeuwenhoek 78, 23–31 (2000). https://doi.org/10.1023/A:1002779201379

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• Issue Date: July 2000
• DOI: https://doi.org/10.1023/A:1002779201379