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In the present study we have analyzed protein oxidation
on Escherichia coli when these cells were submitted
to different stress conditions such as hydrogen peroxide,
superoxide-generating compounds, and iron overloading.
Carbonyl groups on oxidized cell proteins were
examined by Western blot immunoassay. When anaerobically
grown E. coli cells were exposed to hydrogen
peroxide stress, alcohol dehydrogenase E, elongation
factor G, the heat shock protein DNA K, oligopeptidebinding
protein A, enolase, and the outer membrane
protein A were identified as the major protein targets. A
similar immunostained band pattern was found when
cells were shifted from anaerobic to aerobic conditions
in the presence of different concentrations of iron; it is
relevant to note that oxidation of outer membrane protein
C, not observed in peroxide stress conditions, was
clearly detected as the concentration of iron was increased
in the culture media. The hydrogen peroxide
stress performed under aerobic conditions affected the
b-subunit of F0F1-ATPase; the rest of the oxidized protein
pattern was very similar to that found for anaerobic
conditions, with the exception of alcohol dehydrogenase
E, a protein not synthesized aerobically. Cells submitted
to superoxide stress using menadione showed a more
specific pattern in which elongation factor G and the
b-subunit of F0F1-ATPase were affected significantly.
When paraquat was used, although the degree of oxidative
damage was lower, the same two modified proteins
were detected, and DNA K was also clearly damaged.
Cell viability was affected to different extents depending
on the type of stress exerted. The results described
in this paper provide data about the in vivo effects of
oxidative stress on protein oxidation and give insights
into understanding how such modifications can affect
cellular functions.
This work was supported in part by Grant PB94-0829-C02-02 from the Dirección General de Investigación Científica y Técnica and the Comissionat per Universitats i Recerca de la Generalitat de Catalunya and Ajuntament of Lleida (Spain). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. |
