Formation of 8-oxoguanine in cellular DNA of Escherichia coli strains defective in different antioxidant defences.
This paper examines the relationship in Escherichia coli between the in vivo content of 8-oxoguanine (8-oxoG) in chromosomal DNA and deficiencies of various key antioxidant defences. The structural genes for catalases (katG and katE), cytosolic superoxide dismutases (sodA and sodB) or formamidopyrimidine-DNA glycosylase (fpg) were inactivated to obtain bacterial strains lacking the scavenger enzymes for H2O2 or O2.- or the DNA repair protein for 8-oxoG. Wild-type bacteria showed 5-fold increased sensitivity to both lethality and mutagenesis by H2O2 in K medium (1% casamino acids and 1% glucose), as compared with nutrient broth. This higher sensitivity was associated with increased chromosomal oxidative damage, estimated as the 8-oxodG content, and with a marked decrease in both catalase and SOD activities. Bacteria lacking both cytosolic SODs (sodA sodB mutant) displayed increased 8-oxodG content in chromosomal DNA (2.8-fold that of the wild-type) when grown under standard aerated conditions. Comparatively, no significant difference in 8-oxodG content was observed in cells grown without aeration. Bacteria totally devoid of catalase activity (katG katE mutant) showed wild-type contents of 8-oxodG in chromosomal DNA when grown under aerated conditions. Nevertheless, the protective role of catalase in preventing formation of 8-oxodG in chromosomal DNA became evident under oxidative stress conditions: growth under hyperoxygenation and, particularly, following H2O2 exposure. Catalase deficiency resulted in a dramatic decrease in viability after H2O2 exposure. A deficiency of Fpg protein also sensitized E.coli to H2O2 lethality, though to lesser extent than a deficiency of catalase activity. However, the scavenger enzyme and the DNA repair protein protected equally against 8-oxoG formed in vivo upon H2O2 treatment.
Alhama
J
J
Departamento de Bioquímica y Biología Molecular, Universidad de Córdoba, España.
Ruiz-Laguna
J
J
Rodriguez-Ariza
A
A
Toribio
F
F
López-Barea
J
J
Pueyo
C
C
D000469:Alkaline Phosphatase / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002374:Catalase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D003470:Culture Media; D004269:DNA, Bacterial / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D004926:Escherichia coli / Q000145:classification / Q000187:drug effects / Q000235:genetics*; D006147:Guanine / Q000031:analogs & derivatives* / Q000096:biosynthesis; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006868:Hydrolysis; D016296:Mutagenesis; D016877:Oxidants / Q000378:metabolism; D018384:Oxidative Stress; D010544:Peroxidases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D013482:Superoxide Dismutase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D004269:DNA, Bacterial; D016877:Oxidants; C024829:8-hydroxyguanine; D006147:Guanine; D006861:Hydrogen Peroxide; D010544:Peroxidases; C102664:hydroperoxidase II; D002374:Catalase; C076136:catalase HPI; C095500:SodA protein, Bacteria; C087504:SodB protein, Bacteria; D013482:Superoxide Dismutase; D000469:Alkaline Phosphatase