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Molecular Genetics and Genomics

, Volume 269, Issue 5, pp 640–648 | Cite as

Patterns of protein carbonylation following oxidative stress in wild-type and sigB Bacillus subtilis cells

  • J. Mostertz
  • M. HeckerEmail author
Original Paper

Abstract

Oxidative stress causes damage to nucleic acids, membrane lipids and proteins. One striking effect is the metal-catalyzed, site-specific carbonylation of proteins. In the gram-positive soil bacterium Bacillus subtilis, the PerR-dependent specific stress response and the σB-dependent general stress response act together to make cells more resistant to oxidative stress. In this study, we analyzed the carbonylation of cytoplasmic proteins in response to hydrogen peroxide stress in B. subtilis. Furthermore, we asked whether the σB-dependent response to oxidative stress also confers protection against protein carbonylation. To monitor the amount and specificity of protein damage, carbonyls were derivatized with 2,4-dinitrophenylhydrazine, and the resulting stable hydrazones were detected by immunoanalysis of proteins separated by one- or two-dimensional gel electrophoresis. The overall level of protein carbonylation increased strongly in cells treated with hydrogen peroxide. Several proteins, including the elongation factors EF-G, TufA and EF-Ts, were found to be highly carbonylated. Induction of the peroxide specific stress response by treatment with sub-lethal peroxide concentrations, prior to exposure to otherwise lethal levels of peroxide, markedly reduced the degree of protein carbonylation. Cells starved for glucose also showed only minor amounts of peroxide-mediated protein carbonylation compared to exponentially growing cells. We could not detect any differences between wild-type and ΔsigB cells starved for glucose or preadapted by heat treatment with respect to the amount or specificity of protein damage incurred upon subsequent exposure to peroxide stress. However, artificial preloading with proteins that are normally induced by σB-dependent mechanisms resulted in a lower level of protein carbonylation when cells were later subjected to oxidative stress.

Keywords

General stress response Oxidative stress Protein carbonylation Cross adaptation  sigB mutant 

Notes

Acknowledgements

We thank Gerhard Mittenhuber and Finn Viehberg for carefully reading the manuscript. Birgit Voigt, Haike Henkel and Björn Maul are gratefully acknowledged for providing plasmids for antibody generation, purified GsiB and anti-GsiB antibody. This work was supported by grants from the Deutsche Forschungsgemeinschaft, the BMBF, the Bildungsministerium Land Mecklenburg-Vorpommern and the Fonds der Chemischen Industrie to M.H.

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Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  1. 1.Institut für MikrobiologieErnst-Moritz-Arndt-Universität GreifswaldGreifswaldGermany

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