Endogenous Superoxide Dismutase Levels Regulate Iron-Dependent Hydroxyl Radical Formation in Escherichia coli Exposed to Hydrogen Peroxide
01 Feb 1998-Journal of Bacteriology (American Society for Microbiology)-Vol. 180, Iss: 3, pp 622-625
TL;DR: The hypothesis that a resulting increase in .OH formation generated by Fenton chemistry is responsible for the observed enhancement of DNA damage and the increased susceptibility to H2O2-mediated killing seen in these mutants lacking SOD is supported.
Abstract: Aerobic organisms contain antioxidant enzymes, such as superoxide dismutase (SOD) and catalase, to protect them from both direct and indirect effects of reactive oxygen species, such as O2.- and H2O2. Previous work by others has shown that Escherichia coli mutants lacking SOD not only are more susceptible to DNA damage and killing by H2O2 but also contain larger pools of intracellular free iron. The present study investigated if SOD-deficient E. coli cells are exposed to increased levels of hydroxyl radical (.OH) as a consequence of the reaction of H2O2 with this increased iron pool. When the parental E. coli strain AB1157 was exposed to H2O2 in the presence of an alpha-(4-pyridyl-1-oxide)-N-tert-butyl-nitrone (4-POBN)-ethanol spin-trapping system, the 4-POBN-.CH(CH3)OH spin adduct was detectable by electron paramagnetic resonance (EPR) spectroscopy, indicating .OH production. When the isogenic E. coli mutant JI132, lacking both Fe- and Mn-containing SODs, was exposed to H2O2 in a similar manner, the magnitude of .OH spin trapped was significantly greater than with the control strain. Preincubation of the bacteria with the iron chelator deferoxamine markedly inhibited the magnitude of .OH spin trapped. Exogenous SOD failed to inhibit .OH formation, indicating the need for intracellular SOD. Redox-active iron, defined as EPR-detectable ascorbyl radical, was greater in the SOD-deficient strain than in the control strain. These studies (i) extend recent data from others demonstrating increased levels of iron in E. coli SOD mutants and (ii) support the hypothesis that a resulting increase in .OH formation generated by Fenton chemistry is responsible for the observed enhancement of DNA damage and the increased susceptibility to H2O2-mediated killing seen in these mutants lacking SOD.
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TL;DR: Lignin was isolated from corn stover degraded by Irpex lacteus CD2, and the structure alterations were analyzed by elemental analysis, FTIR, 13C NMR, 1H NMR and UV spectra as discussed by the authors.
50 citations
TL;DR: This review focuses on specific molecular pathways that allow V. fischeri to establish this exquisite bacteria–host interaction and manages these environments to outcompete all other bacterial species for colonization of E. scolopes.
Abstract: Bacteria successfully colonize distinct niches because they can sense and appropriately respond to a variety of environmental signals. Of particular interest is how a bacterium negotiates the multiple, complex environments posed during successful infection of an animal host. One tractable model system to study how a bacterium manages a host’s multiple environments is the symbiotic relationship between the marine bacterium, Vibrio fischeri, and its squid host, Euprymna scolopes. V. fischeri encounters many different host surroundings ranging from initial contact with the squid to ultimate colonization of a specialized organ known as the light organ. For example, upon recognition of the squid, V. fischeri forms a biofilm aggregate outside the light organ that is required for efficient colonization. The bacteria then disperse from this biofilm to enter the organ, where they are exposed to nitric oxide, a molecule that can act as both a signal and an antimicrobial. After successfully managing this potentially hostile environment, V. fischeri finally establish their niche in the deep crypts of the light organ where the bacteria bioluminesce in a pheromone-dependent fashion, a phenotype that E. scolopes utilizes for anti-predation purposes. The mechanism by which V. fischeri manages these environments to outcompete all other bacterial species for colonization of E. scolopes is an important and intriguing question that will permit valuable insights into how a bacterium successfully associates with a host. This review focuses on specific molecular pathways that allow V. fischeri to establish this exquisite bacteria-host interaction.
48 citations
Cites background from "Endogenous Superoxide Dismutase Lev..."
...…NO sensing through H-NOX “primes” V. fischeri for the crypt environment, where survival may depend upon the ability of the bacterium to combat the formation of these hydroxyl radicals by controlling the levels of free iron (McCormick et al., 1998; Semsey et al., 2006; Wang et al., 2010a; Figure 5)....
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TL;DR: Results showed that the hemoglobin-expressing strain of Enterobacter aerogenes engineered to produce the Vitreoscilla Hb is quite distinct in terms of growth/survival properties and activity of antioxidant enzymes from that of non-hemoglobin counterparts.
44 citations
TL;DR: The ferritin was particularly important for the bacterium to survive under iron-depleted conditions (both haemin and transferrin starvation), indicating that intracellular iron is stored in Ferritin regardless of the iron source and that the iron stored inFerritin is utilized under iron -restricted conditions.
Abstract: Porphyromonas gingivalis is an obligate anaerobe that utilizes haem, transferrin and haemoglobin efficiently as sources of iron for growth, and has the ability to store haem on its cell surface, resulting in black pigmentation of colonies on blood agar plates. However, little is known about intracellular iron storage in this organism. Ferritin is one of the intracellular iron-storage proteins and may also contribute to the protection of organisms against oxidative stresses generated by intracellular free iron. A ferritin-like protein was purified from P. gingivalis and the encoding gene (ftn) was cloned from chromosomal DNA using information on its amino-terminal amino acid sequence. Comparison of the amino acid sequence deduced from the nucleotide sequence of ftn with those of known ferritins and bacterioferritins identified the protein as a ferritin and positioned it between proteins from the Proteobacteria and Thermotogales. The P. gingivalis ferritin was found to contain non-haem iron, thus confirming its identity. Construction and characterization of a P. gingivalis ferritin-deficient mutant revealed that the ferritin was particularly important for the bacterium to survive under iron-depleted conditions (both haemin and transferrin starvation), indicating that intracellular iron is stored in ferritin regardless of the iron source and that the iron stored in ferritin is utilized under iron-restricted conditions. However, the ferritin appeared not to contribute to protection against oxidative stresses caused by peroxides and atmospheric oxygen.
42 citations
Cites background from "Endogenous Superoxide Dismutase Lev..."
...Free iron has the ability to form a highly reactive oxidant, the hydroxyl radical, through the Fenton reaction, which can cause damage to cellular components including DNA and membranes (AbdulTehrani, 1999; McCormick et al., 1998; Keyer et al., 1995; O’Connell et al., 1985)....
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...Recent studies using E. coli have demonstrated that superoxide toxicity is mainly due to its role in accelerating the Fenton reaction (Keyer & Imlay, 1996; McCormick et al., 1998)....
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...coli have demonstrated that superoxide toxicity is mainly due to its role in accelerating the Fenton reaction (Keyer & Imlay, 1996; McCormick et al., 1998)....
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TL;DR: The results suggest that dpr and sod in S. mutans are involved in coexistence with S. sanguinis, and PerR is associated with resistance to H2O2 in regulating the expression of Dpr.
Abstract: Large numbers of bacteria coexist in the oral cavity. Streptococcus sanguinis, one of the major bacteria in dental plaque, produces hydrogen peroxide (H2O2), which interferes with the growth of other bacteria. Streptococcus mutans, a cariogenic bacterium, can coexist with S. sanguinis in dental plaque, but to do so, it needs a means of detoxifying the H2O2 produced by S. sanguinis. In this study, we investigated the association of three oxidative stress factors, Dpr, superoxide dismutase (SOD), and AhpCF, with the resistance of S. sanguinis to H2O2. The knockout of dpr and sod significantly increased susceptibility to H2O2, while the knockout of ahpCF had no apparent effect on susceptibility. In particular, dpr inactivation resulted in hypersensitivity to H2O2. Next, we sought to identify the factor(s) involved in the regulation of these oxidative stress genes and found that PerR negatively regulated dpr expression. The knockout of perR caused increased dpr expression levels, resulting in low-level susceptibility to H2O2 compared with the wild type. Furthermore, we evaluated the roles of perR, dpr, and sod when S. mutans was cocultured with S. sanguinis. Culturing of the dpr or sod mutant with S. sanguinis showed a significant decrease in the S. mutans population ratio compared with the wild type, while the perR mutant increased the ratio. Our results suggest that dpr and sod in S. mutans are involved in coexistence with S. sanguinis, and PerR is associated with resistance to H2O2 in regulating the expression of Dpr.
42 citations
Cites background from "Endogenous Superoxide Dismutase Lev..."
...By the addition of H2O2 to the medium, the accumulated O2 in the sod mutant reacts with H2O2, causing the formation of OH, which is considered to be highly toxic to bacteria (35)....
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References
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Book•
13 Jun 1985
TL;DR: 1. Oxygen is a toxic gas - an introduction to oxygen toxicity and reactive species, and the chemistry of free radicals and related 'reactive species'
Abstract: 1. Oxygen is a toxic gas - an introductionto oxygen toxicity and reactive species 2. The chemistry of free radicals and related 'reactive species' 3. Antioxidant defences Endogenous and Diet Derived 4. Cellular responses to oxidative stress: adaptation, damage, repair, senescence and death 5. Measurement of reactive species 6. Reactive species can pose special problems needing special solutions. Some examples. 7. Reactive species can be useful some more examples 8. Reactive species can be poisonous: their role in toxicology 9. Reactive species and disease: fact, fiction or filibuster? 10. Ageing, nutrition, disease, and therapy: A role for antioxidants?
21,528 citations
"Endogenous Superoxide Dismutase Lev..." refers background in this paper
...rapidly reacts with itself (dismutes) to form H2O2 (7)....
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...Although the aerobic metabolism of bacteria optimally results in the near simultaneous four-electron reduction of O2 to H2O, a variable percentage of O2 reduction occurs initially via either one-electron reduction of O2 to superoxide (O2 ) or divalent reduction to H2O2 (7)....
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TL;DR: It is proposed that the cell may also decrease such toxicity by diminishing available NAD(P)H and by utilizing oxygen itself to scavenge active free radicals into superoxide, which is then destroyed by superoxide dismutase.
Abstract: A major portion of the toxicity of hydrogen peroxide in Escherichia coli is attributed to DNA damage mediated by a Fenton reaction that generates active forms of hydroxyl radicals from hydrogen peroxide, DNA-bound iron, and a constant source of reducing equivalents. Kinetic peculiarities of DNA damage production by hydrogen peroxide in vivo can be reproduced by including DNA in an in vitro Fenton reaction system in which iron catalyzes the univalent reduction of hydrogen peroxide by the reduced form of nicotinamide adenine dinucleotide (NADH). To minimize the toxicity of oxygen radicals, the cell utilizes scavengers of these radicals and DNA repair enzymes. On the basis of observations with the model system, it is proposed that the cell may also decrease such toxicity by diminishing available NAD(P)H and by utilizing oxygen itself to scavenge active free radicals into superoxide, which is then destroyed by superoxide dismutase.
1,997 citations
"Endogenous Superoxide Dismutase Lev..." refers background in this paper
...Pretreatment of the JI132 (SOD-deficient) bacteria with DFO greatly reduced the magnitude of OH generation, confirming that it arose as a consequence of Fenton chemistry, as iron bound to DFO is no longer available for this redox chemistry (10)....
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TL;DR: Aerotolerant anaerobes, which survive exposure to air and metabolize oxygen to a limited extent but do not contain cytochrome systems, were found to be devoid of catalase activity but did exhibit superoxide dismutase activity.
Abstract: The distribution of catalase and superoxide dismutase has been examined in various micro-organisms. Strict anaerobes exhibited no superoxide dismutase and, generally, no catalase activity. All aerobic organisms containing cytochrome systems were found to contain both superoxide dismutase and catalase. Aerotolerant anaerobes, which survive exposure to air and metabolize oxygen to a limited extent but do not contain cytochrome systems, were found to be devoid of catalase activity but did exhibit superoxide dismutase activity. This distribution is consistent with the proposal that the prime physiological function of superoxide dismutase is protection of oxygen-metabolizing organisms against the potentially detrimental effects of the superoxide free radical, a biologically produced intermediate resulting from the univalent reduction of molecular oxygen.
974 citations
"Endogenous Superoxide Dismutase Lev..." refers background in this paper
...Most bacteria, including Escherichia coli, contain superoxide dismutase (SOD) and catalase as means of eliminating O2 z2 and H2O2, respectively (16, 17)....
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TL;DR: This presentation discusses the role of catalytic metals in free radical-mediated oxidations, ascorbate as both a pro-oxidant and an antioxidant, use of asCorbate to determine adventitious catalytic metal concentrations, and uses of ascorBate radical as a marker of oxidative stress.
Abstract: Trace levels of transition metals can participate in the metal-catalyzed Haber-Weiss reaction (superoxide-driven Fenton reaction) as well as catalyze the oxidation of ascorbate. Generally ascorbate is thought of as an excellent reducing agent; it is able to serve as a donor antioxidant in free radical-mediated oxidation processes. However, as a reducing agent it is also able to reduce redox-active metals such as copper and iron, thereby increasing the pro-oxidant chemistry of these metals. Thus ascorbate can serve as both a pro-oxidant and an antioxidant. In general, at low ascorbate concentrations, ascorbate is prone to be a pro-oxidant, and at high concentrations, it will tend to be an antioxidant. Hence there is a crossover effect. We propose that the "position" of this crossover effect is a function of the catalytic metal concentration. In this presentation, we discuss: (1) the role of catalytic metals in free radical-mediated oxidations; (2) ascorbate as both a pro-oxidant and an antioxidant; (3) catalytic metal catalysis of ascorbate oxidation; (4) use of ascorbate to determine adventitious catalytic metal concentrations; (5) use of ascorbate radical as a marker of oxidative stress; and (6) use of ascorbate and iron as free radical pro-oxidants in photodynamic therapy of cancer.
851 citations
TL;DR: In this article, the authors show that the level of loose iron in severely superoxide-stressed cells greatly exceeds that of unstressed cells, and that both growth defects and DNA damage caused by superoxide ensue from its ability to damage a subset of iron-sulfur clusters.
Abstract: Superoxide promotes hydroxyl-radical formation and consequent DNA
damage in cells of all types. The long-standing hypothesis that it
primarily does so by delivering electrons to adventitious iron on DNA
was refuted by recent studies in Escherichia coli.
Alternative proposals have suggested that superoxide may accelerate
oxidative DNA damage by leaching iron from storage proteins or enzymic
[4Fe-4S] clusters. The released iron might then deposit on the
surface of the DNA, where it could catalyze the formation of DNA
oxidants using other electron donors. The latter model is affirmed by
the experiments described here. Whole-cell electron paramagnetic
resonance demonstrated that the level of loose iron in
superoxide-stressed cells greatly exceeds that of unstressed cells.
Bacterial iron storage proteins were not the major source for free
iron, since superoxide also increased iron levels in mutants lacking
these iron storage proteins. However, overproduction of an enzyme
containing a labile [4Fe-4S] cluster dramatically increased the free
iron content of cells when they were growing in air. The rates of
spontaneous mutagenesis and DNA damage from exogenous
H2O2 increased commensurately. It is striking
that both growth defects and DNA damage caused by superoxide ensue from
its ability to damage a subset of iron–sulfur clusters.
803 citations