Effects of Cystine and Hydrogen Peroxide on Glutathione Status and Expression of Antioxidant Genes in Escherichia coli

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G. V. Smirnova^*, N. G. Muzyka, and O. N. Oktyabrsky

Institute of Ecology and Genetics of Microorganisms, Russian Academy of Sciences, ul. Goleva 13, 614081 Perm, Russia; fax: (3422) 446-711; E-mail: smirnova@iegm.ru

^* To whom correspondence should be addressed.

Received May 25, 2004; Revision received July 20, 2004

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Cysteine or cystine was earlier shown to multiply enhance the toxic effect of hydrogen peroxide on Escherichia coli cells. In the present work, the treatment of E. coli with H₂O₂ in the presence of cystine increased fivefold the level of extracellular oxidized glutathione (GSSG_out) and decreased fivefold the GSH/GSSG_out ratio (from 16.8 to 3.6). The same treatment of cells with deficiency in glutathione oxidoreductase (GOR) resulted in even more severe oxidation of GSH_out, so that the level of oxidized glutathione exceeded that of reduced glutathione and the GSH/GSSG_out ratio decreased to 0.4. Addition of cystine to the GOR deficient cells resulted in significant oxidation of extracellular glutathione even in the absence of oxidant and in tenfold increase in intracellular oxidized glutathione along with a decrease in the GSH/GSSG_out ratio from 282 to 26. However, in the cytoplasm of wild type cells, the level of oxidized glutathione (GSSG_in) was changed insignificantly and the GSH/GSSG_in ratio increased by 26% (from 330 to 415). Data on glutathione status and cystine reduction in the E. coli gsh and gor mutants suggested that exogenous cystine at first should be reduced with extracellular GSH outside the cells and then imported into them. The high toxicity of H₂O₂ in the presence of cystine resulted in disorders of membrane functions and inhibition of the expression of genes including those responsible for neutralization of oxidants and DNA repair.

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KEY WORDS: hydrogen peroxide, cystine, gene expression, reduced and oxidized glutathione, Escherichia coli

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