Sensitivity of various Escherichia coli strains to 2,4,6-trinitrotoluene

Kurinenko, B. M.; Denivarova, N. A.; Yakovleva, Galina

doi:10.1007/s10438-005-0009-5

Cited by 4 publications

(5 citation statements)

References 5 publications

(10 reference statements)

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“…Also, several reports have described the reduction of TNT by E. coli but not its denitration (6,14,22,23). Only one recent study has mentioned denitration of TNT by E. coli, but no quantitative data were provided and TNT was not the sole nitrogen source (13). The objectives of this study were to determine the kinetics of TNT denitration by E. coli and identify TNT denitrated metabolites.…”

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confidence: 99%

Aerobic Growth of Escherichia coli with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts

Stenuit¹,

Eyers²,

Rozenberg

et al. 2006

Appl Environ Microbiol

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Escherichia coli grew aerobically with 2,4,6-trinitrotoluene (TNT) as sole nitrogen source and caused TNT's partial denitration. This reaction was enhanced in nongrowing cell suspensions with 0.516 mol nitrite released per mol TNT. Cell extracts denitrated TNT in the presence of NAD(P)H. Isomers of amino-dimethyl-tetranitrobiphenyl were detected and confirmed with U-15 N-labeled TNT.2,4,6-Trinitrotoluene (TNT) is recalcitrant to microbial degradation. Denitration (defined as the release of nitrite) is a critical step for further mineralization of TNT (19). A welldescribed TNT denitration pathway involves a nucleophilic addition of hydride ions to the aromatic ring with subsequent nitrite release. Three enzymes performing this addition in the presence of NAD(P)H have been characterized so far: pentaerythritol tetranitrate reductase of Enterobacter cloacae PB2 (9), xenobiotic reductase B (XenB) of Pseudomonas fluorescens I-C (15), and N-ethylmaleimide (NEM) reductase of Escherichia coli (21). In vitro denitration of TNT with purified NEM reductase was described by Williams et al. (21), but the authors did not provide quantitative data with E. coli cells. Also, several reports have described the reduction of TNT by E. coli but not its denitration (6,14,22,23). Only one recent study has mentioned denitration of TNT by E. coli, but no quantitative data were provided and TNT was not the sole nitrogen source (13). The objectives of this study were to determine the kinetics of TNT denitration by E. coli and identify TNT denitrated metabolites.E. coli strains EPI300 (Epicentre Technologies, Madison, WI) and LK111 (24) were routinely cultivated at 37°C in LuriaBertani (LB) broth. Cells were harvested at mid-exponential phase and washed three times with phosphate-buffered saline (containing, per liter, 7 g of Na 2 HPO 4 ·12H 2 O, 3 g of KH 2 PO 4 , 1 g of NaCl). Cells were resuspended in 20 ml of phosphatebuffered saline and used for TNT biodegradation assays.TNT was obtained from Nobel Explosives (Châtelet, Belgium) and was 99.5% pure by high-performance liquid chromatography. Growing cell experiments were carried out in modified mineral salts medium (10) containing 20 mM of glycerol or glucose and TNT as the sole nitrogen source. The medium was inoculated at an optical density at 600 nm (OD 600 ) of 0.025 and incubated at 37°C and 250 rpm. Controls consisted of flasks without TNT, flasks without cells, flasks without a carbon source, and flasks with 200 ppm of Hg 2 Cl 2 and without a carbon source. Nitrite, TNT, and metabolites were quantitatively determined as previously described (7).Bacterial growth of E. coli EPI300 was observed with glycerol and TNT (606 M on the basis of high-performance liquid chromatography analysis) as the sole nitrogen source (Fig. 1A). The growth was relatively fast over the first 26 h, reaching a plateau of 0.120 OD 600 units after 117 h of incubation. With glucose and TNT (588 M), the bacterial growth profile was similar but the OD 600 reached 0.320 after 117 h (Fig. 1B). Without TNT, no si...

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confidence: 99%

Aerobic Growth of Escherichia coli with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts

Stenuit¹,

Eyers²,

Rozenberg

et al. 2006

Appl Environ Microbiol

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“…The efficiency of microbial remediation of NACs is constrained by two major factors: (1) low rate of degradation hampering time-efficient clean-up of contaminants (particularly at high loading rates); and (2) the relatively low tolerance of microbes to chemical toxicity [ 3 ]. Studies on the E. coli TNT tolerance have reported various multiple thresholds ranging from 66 to 200 mg/L [ 31 , 32 ]. However, there is meager information on how variation of initial TNT concentration within the nontoxic range may affect degradation efficiency.…”

Section: Resultsmentioning

confidence: 99%

“…However, there is meager information on how variation of initial TNT concentration within the nontoxic range may affect degradation efficiency. To gain insight into this, we conducted DFBA at various initial TNT concentrations up to the maximum toxic threshold [ 31 ], under both aerobic and anaerobic regimes. As shown in Figure 10 , lower TNT concentrations are correlated with higher productivity.…”

Section: Resultsmentioning

confidence: 99%

“…Also at lower concentrations productivity is higher in anaerobic vs. aerobic processes. On the other hands, as concentration exceeds 0.8 mg/mL (typical values in batch cultivation [ 27 , 31 ]), productivity tend to lose negative sensitivity to both concentration and aeration mode, approaching a constant level. Low sensitivity of productivity to concentration is advantageous because it permits increased biotransformation yield (by operating at high TNT concentration) with limited negative effect on productivity.…”

Section: Resultsmentioning

confidence: 99%

“…E. coli is able to derive assimilatory ammonium from TNT, placing it among the limited number of microbes capable of growing on TNT as the sole nitrogen source [ 27 , 28 ]. In addition, the organism has shown significant tolerance against TNT toxicity while maintaining the biotransformation capability at relatively high TNT concentrations [ 31 , 32 ]. These characteristic advantages together with the wealth of information on physiology and molecular biology of E. coli , highlight this organism as a promising biocatalyst for TNT bioremediation processes.…”

Section: Introductionmentioning

confidence: 99%

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Systems Biology Approach to Bioremediation of Nitroaromatics: Constraint-Based Analysis of 2,4,6-Trinitrotoluene Biotransformation by Escherichia coli

et al. 2017

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Microbial remediation of nitroaromatic compounds (NACs) is a promising environmentally friendly and cost-effective approach to the removal of these life-threating agents. Escherichia coli (E. coli) has shown remarkable capability for the biotransformation of 2,4,6-trinitro-toluene (TNT). Efforts to develop E. coli as an efficient TNT degrading biocatalyst will benefit from holistic flux-level description of interactions between multiple TNT transforming pathways operating in the strain. To gain such an insight, we extended the genome-scale constraint-based model of E. coli to account for a curated version of major TNT transformation pathways known or evidently hypothesized to be active in E. coli in present of TNT. Using constraint-based analysis (CBA) methods, we then performed several series of in silico experiments to elucidate the contribution of these pathways individually or in combination to the E. coli TNT transformation capacity. Results of our analyses were validated by replicating several experimentally observed TNT degradation phenotypes in E. coli cultures. We further used the extended model to explore the influence of process parameters, including aeration regime, TNT concentration, cell density, and carbon source on TNT degradation efficiency. We also conducted an in silico metabolic engineering study to design a series of E. coli mutants capable of degrading TNT at higher yield compared with the wild-type strain. Our study, therefore, extends the application of CBA to bioremediation of nitroaromatics and demonstrates the usefulness of this approach to inform bioremediation research.

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TNT biotransformation: when chemistry confronts mineralization

Smets

Yin²,

Esteve‐Núñez

2007

Appl Microbiol Biotechnol

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Our understanding of the genetics and biochemistry of microbial 2,4,6-trinitrotoluene (TNT) biotransformation has advanced significantly during the past 10 years, and biotreatment technologies have developed. In this review, we summarize this new knowledge. A number of enzyme classes involved in TNT biotransformation include the type I nitroreductases, the old yellow enzyme family, a respiration-associated nitroreductase, and possibly ring hydroxylating dioxygenases. Several strains harbor dual pathways: nitroreduction (reduction of the nitro group in TNT to a hydroxylamino and/or amino group) and denitration (reduction of the aromatic ring of TNT to Meisenheimer complexes with nitrite release). TNT can serve as a nitrogen source for some strains, and the postulated mechanism involves ammonia release from hydroxylamino intermediates. Field biotreatment technologies indicate that both stimulation of microbial nitroreduction and phytoremediation result in significant and permanent immobilization of TNT via its metabolites. While the possibility for TNT mineralization was rekindled with the discovery of TNT denitration and oxygenolytic and respiration-associated pathways, further characterization of responsible enzymes and their reaction mechanisms are required.

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Sensitivity of various Escherichia coli strains to 2,4,6-trinitrotoluene

Cited by 4 publications

References 5 publications

Aerobic Growth of Escherichia coli with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts

Aerobic Growth of Escherichia coli with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts

Systems Biology Approach to Bioremediation of Nitroaromatics: Constraint-Based Analysis of 2,4,6-Trinitrotoluene Biotransformation by Escherichia coli

TNT biotransformation: when chemistry confronts mineralization

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