Characteristics of major<i>Escherichia coli</i>reductases involved in aerobic nitro and azo reduction

Mercier, Corinne; Chalansonnet, Valérie; Orenga, Sylvain; Gilbert, Christophe

doi:10.1111/jam.12294

Cited by 32 publications

(37 citation statements)

References 41 publications

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“…This result was also consistent with recent findings, which demonstrated the prominent role of E. coli AzoR-purified enzyme under aerobic conditions (Mercier et al 2013). The remarkably high rate of decolorization for B46 was found to be dependent on the amount of biomass and the concentration of dye.…”

Section: Discussionsupporting

confidence: 82%

Decolorization of acid and basic dyes: understanding the metabolic degradation and cell-induced adsorption/precipitation by Escherichia coli

Cerboneschi

Corsi

Bianchini

et al. 2015

Appl Microbiol Biotechnol

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Escherichia coli strain DH5α was successfully employed in the decolorization of commercial anthraquinone and azo dyes, belonging to the general classes of acid or basic dyes. The bacteria showed an aptitude to survive at different pH values on any dye solution tested, and a rapid decolorization was obtained under aerobic conditions for the whole collection of dyes. A deep investigation about the mode of action of E. coli was carried out to demonstrate that dye decolorization mainly occurred via three different pathways, specifically bacterial induced precipitation, cell wall adsorption, and metabolism, whose weight was correlated with the chemical nature of the dye. In the case of basic azo dyes, an unexpected fast decolorization was observed after just 2-h postinoculation under aerobic conditions, suggesting that metabolism was the main mechanism involved in basic azo dye degradation, as unequivocally demonstrated by mass spectrometric analysis. The reductive cleavage of the azo group by E. coli on basic azo dyes was also further demonstrated by the inhibition of decolorization occurring when glucose was added to the dye solution. Moreover, no residual toxicity was found in the E. coli-treated basic azo dye solutions by performing Daphnia magna acute toxicity assays. The results of the present study demonstrated that E. coli can be simply exploited for its natural metabolic pathways, without applying any recombinant technology. The high versatility and adaptability of this bacterium could encourage its involvement in industrial bioremediation of textile and leather dyeing wastewaters.

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Section: Discussionsupporting

confidence: 82%

Decolorization of acid and basic dyes: understanding the metabolic degradation and cell-induced adsorption/precipitation by Escherichia coli

Cerboneschi

Corsi

Bianchini

et al. 2015

Appl Microbiol Biotechnol

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“…These results corroborate a previous study (7) and can probably be attributed to the oxygensensitive nitroreductase system, which becomes active in the absence of oxygen and hence compensates for the (partial) loss of activity of the oxygen-insensitive nitroreductase system (19,20). In addition, another study using single-and double-knockout E. coli mutants targeting nfsA and nfsB showed that, at low oxygen levels, the mutants were still able to reduce the nitro compound 7-nitrocoumarin-3-carboxylic acid (7NCCA), indicating that NfsA and NfsB are not required for nitroreduction under anaerobic growth conditions (21). In ST2747-an, mutations in nfsA or nfsB could not be detected, although a 2-fold MIC increase was observed.…”

Section: Discussionmentioning

confidence: 99%

“…FMN is tightly associated with NfsA and NfsB, where it serves as a mediator for electron transfer from NAD(P)H to various substrates for nitroreduction (24). En- zymatic assays with 7NCCA demonstrated that both nitroreductases were greatly dependent on FMN for nitroreduction to proceed (21). The 12-nucleotide deletion in ribE resulted in a four-amino acid loss (TKAG) in the active site of lumazine synthase, at positions 131 to 134 (Fig.…”

Section: Discussionmentioning

confidence: 99%

An In Vitro Deletion in ribE Encoding Lumazine Synthase Contributes to Nitrofurantoin Resistance in Escherichia coli

Vervoort

Xavier

Stewardson

et al. 2014

Antimicrob Agents Chemother

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e Nitrofurantoin has been used for decades for the treatment of urinary tract infections (UTIs), but clinically significant resistance in Escherichia coli is uncommon. Nitrofurantoin concentrations in the gastrointestinal tract tend to be low, which might facilitate selection of nitrofurantoin-resistant (NIT-R) strains in the gut flora. We subjected two nitrofurantoin-susceptible intestinal E. coli strains (ST540-p and ST2747-p) to increasing nitrofurantoin concentrations under aerobic and anaerobic conditions. Whole-genome sequencing was performed for both susceptible isolates and selected mutants that exhibited the highest nitrofurantoin resistance levels aerobically (ST540-a and ST2747-a) and anaerobically (ST540-an and ST2747-an). ST540-a/ST540-an and ST2747-a (aerobic MICs of >64 g/ml) harbored mutations in the known nitrofurantoin resistance determinants nfsA and/or nfsB, which encode oxygen-insensitive nitroreductases. ST2747-an showed reduced nitrofurantoin susceptibility (aerobic MIC of 32 g/ml) and exhibited remarkable growth deficits but did not harbor nfsA/nfsB mutations. We identified a 12-nucleotide deletion in ribE, encoding lumazine synthase, an essential enzyme involved in the biosynthesis of flavin mononucleotide (FMN), which is an important cofactor for NfsA and NfsB. Complementing ST2747-an with a functional wild-type lumazine synthase restored nitrofurantoin susceptibility. Six NIT-R E. coli isolates (NRCI-1 to NRCI-6) from stools of UTI patients treated with nitrofurantoin, cefuroxime, or a fluoroquinolone harbored mutations in nfsA and/or nfsB but not ribE. Sequencing of the ribE gene in six intestinal and three urinary E. coli strains showing reduced nitrofurantoin susceptibility (MICs of 16 to 48 g/ml) also did not identify any relevant mutations. NRCI-1, NRCI-2, and NRCI-5 exhibited up to 4-fold higher anaerobic MICs, compared to the mutants generated in vitro, presumably because of additional mutations in oxygen-sensitive nitroreductases.

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“…), which are the two major nitroreductases in E. coli MG1655. As E. coli azoreductase AzoR displays nitroreductase activity [6, 18], a similar BLAST protein search was also performed using AzoR as the reference protein (AAC74494.1. ).…”

Section: Methodsmentioning

confidence: 99%

Identification of Enterococcus faecalis enzymes with azoreductases and/or nitroreductase activity

et al. 2017

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BackgroundNitroreductases, NAD(P)H dependent flavoenzymes, are found in most of bacterial species. Even if Enterococcus faecalis strains seems to present such activity because of their sensitivity to nitrofurans, no enzyme has been described.Nitroreductases were separated of others reductases due to their capacity to reduce nitro compounds. They are further classified based on their preference in cofactor: NADH and/or NADPH. However, recently, azoreductases have been studied for their strong activity on nitro compounds, especially nitro pro-drugs. This result suggests a crossing in azo and nitro reductase activities. For the moment, no nitroreductase was demonstrated to possess azoreductase activity. But due to sequence divergence and activity specificity linked to substrates, activity prediction is not evident and biochemical characterisation remains necessary.Identifying enzymes active on these two classes of compounds: azo and nitro is of interest to consider a common physiological role.ResultsFour putative nitroreductases, EF0404, EF0648, EF0655 and EF1181 from Enterococcus faecalis V583 were overexpressed as his-tagged recombinant proteins in Escherichia coli and purified following a native or a denaturing/renaturing protocol. EF0648, EF0655 and EF1181 showed nitroreductase activity and their cofactor preferences were in agreement with their protein sequence phylogeny. EF0404 showed both nitroreductase and azoreductase activity. Interestingly, the biochemical characteristics (substrate and cofactor specificity) of EF0404 resembled the properties of the known azoreductase AzoA. But its sequence matched within nitroreductase group, the same as EF0648.ConclusionsWe here demonstrate nitroreductase activity of the putative reductases identified in the Enterococcus faecalis V583 genome. We identified the first nitroreductase able to reduce directly an azo compound, while its protein sequence is close to others nitroreductases. Consequently, it highlights the difficulty in classifying these enzymes solely on the basis of protein sequence alignment and hereby the necessity to experimentally demonstrate the activity. The results provide additional data to consider a broader functionality of these reductases.

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Characteristics of majorEscherichia colireductases involved in aerobic nitro and azo reduction

Cited by 32 publications

References 41 publications

Decolorization of acid and basic dyes: understanding the metabolic degradation and cell-induced adsorption/precipitation by Escherichia coli

Decolorization of acid and basic dyes: understanding the metabolic degradation and cell-induced adsorption/precipitation by Escherichia coli

An In Vitro Deletion in ribE Encoding Lumazine Synthase Contributes to Nitrofurantoin Resistance in Escherichia coli

Identification of Enterococcus faecalis enzymes with azoreductases and/or nitroreductase activity

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