Identification of novel members of the bacterial azoreductase family in <i>Pseudomonas aeruginosa</i>

Crescente, Vincenzo; Holland, Sinead; Kashyap, Sapna; Polycarpou, Elena; Sim, Edith; Ryan, Ali

doi:10.1042/bj20150856

Cited by 19 publications

(28 citation statements)

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“…For example, it was recently shown that MdaB, ArsH and YieF from P. aeruginosa can reduce different azo compounds while being part of distantly homologous oxido-reductases families with respect to protein sequence. Interestingly, these proteins were also proven to reduce quinones and nitrofurazone [35]. Consequently, biochemical assays are clearly necessary to corroborate the protein homologies.…”

Section: Discussionmentioning

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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Section: Discussionmentioning

confidence: 99%

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

et al. 2017

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“…To support this, enzymes were identified in P. aeruginosa from four families [ArsH, modulator of drug activity B, tryptophan repressor binding protein A (WrbA) and YieF], which are NAD(P)H quinone oxidoreductases that have been characterized in other bacteria (Ryan et al , ). The enzymes identified in P. aeruginosa were cloned, expressed and characterized, and all, except WrbA (the most distantly related homologue), showed azoreductase activity (Crescente et al , ). This indicates that the azoreductase family is larger than originally thought, and more work is needed to characterize further members.…”

Section: Phylogeny Of Azoreductasesmentioning

confidence: 99%

“…Azoreductases in bacteria have been shown to have 10‐ to 100‐fold greater activity against quinones than against azo substrates (Ryan et al , ; Crescente et al , ). Quinones are highly cytotoxic due to their ability to undergo redox cycling via one or two electron reduction but can also cause alkylation of cellular protein and DNA (Bolton et al , ).…”

Section: Nitrofuran and Other Nitroaromatic Drug Activation By Azoredmentioning

confidence: 99%

“…To date, the majority of studies on azoreductases have been performed on enzymes from aerobic bacteria (Nakanishi et al , ; Crescente et al , ); in contrast, only a single azoreductase from a strict anaerobe has been characterized (Morrison et al , ). Several species of gut bacteria have been identified to have azoreductase activities (Wang et al , ); however, studies on individual azoreductases from enteric bacteria are very limited.…”

Section: Introduction To Azoreductasesmentioning

confidence: 99%

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Azoreductases in drug metabolism

Ryan

2016

British J Pharmacology

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Azoreductases are flavoenzymes that have been characterised in a range of prokaryotes and eukaryotes. Bacterial azoreductases are associated with the activation of two classes of drug, azo drugs for the treatment of inflammatory bowel disease and nitrofuran antibiotics. The mechanism of reduction of azo compounds is presented that requires tautomerisation of the azo compound to a quinoneimine and provides a unifying mechanism for the reduction of azo and quinone substrates by azoreductase. The importance of further work in characterisation of azoreductases from enteric bacteria is highlighted to aid in the development of novel drugs for the treatment of colon related disorders.Human azoreductases are known to play a crucial role in the metabolism of a number of quinone containing cancer chemotherapeutic drugs. The mechanism of hydride transfer to quinones, which is shared not only between eukaryotic and prokaryotic azoreductases but the wider family of NAD(P)H quinone oxidoreductases, is outlined. The importance of common SNPs in human azoreductases is described not only in cancer prognosis but also due to their effects on the efficacy of quinone drug based cancer chemotherapeutic regimens. This highlights the need to screen patients for azoreductase SNPs ahead of treatment with these regimens. Non-approved abbreviations5-ASA: 5-aminosalicylate, AcpD: acyl carrier protein phosphodiesterase, ecAzoR: E. coli azoreductase, FAD: flavin adenine dinucleotide, FMN: Flavin mononucleotide, hNQO1/2: human NAD(P)H quinone oxidoreductase 1 and 2, IBD: inflammatory bowel disease, MdaB: Modulator of drug activity B, NAD(P)H: Nicotinamide adenine dinucleotide (phosphate) reduced, NfsB: Nitrofurazone sensitivity protein B, NRH: (Wu et al., 1997) Introduction to azoreductasesAzoreductases are a group of diverse enzymes found in many bacterial and eukaryotic organisms (Fig 1 (Ryan et al., 2014)). The azoreductases discussed in this review are flavin-dependent (typically FMN) enzymes that are able to reductively cleave compounds containing an azo bond. Azo compounds are defined as those that contain an R 1 -N=N-R 2 group, where R 1 and R 2 are typically aromatic groups. Azoreductases are primarily cytosolic enzymes, however they have been shown to be secreted during exposure of bacteria to azo dyes (Morrison et al., 2015). The physiological role of these enzymes in the bacteria remains unclear, however they are constitutively expressed during growth in vitro (Chen et al., 2005;Wang et al., 2007) which suggests a role in homeostasis. The majority of azoreductase research is focused on their use in bioremediation where they can be used for the treatment of waste water contaminated with azo dyes from the textile and cosmetics industries (Singh et al., 2015). This review is unique in focusing on their role in the activation of several classes of drug.To date the majority of studies on azoreductases have been performed on enzymes from aerobic bacteria (Crescente et al., 2016;Nakanishi et al., 2001), in contrast only a single ...

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“…Bioremediation uses plants and microorganisms to degrade which in turn is non-hazardous and eco-friendly. Many species of Acinetobacter, Alcaligenes, Arthrobacter, Aspergillus, Bacillus, Candida, Ewingella, Pseudomonas, Ralstonia, Rhodococcus, Sphingomonas and Streptomyces have been used in bioremediation [1,[5][6][7][8][9][10][11].…”

Section: Introductionmentioning

confidence: 99%

Untitled

2018

Bioinformation

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Cresol is an organic pollutant discharged by pharmaceutical, pesticide, coal and gasification industries causing severe organ failure in humans. Therefore, it is of interest to isolate microbes from contaminated site for degrading cresol. We isolated a strain (CR-13) that survives at 5000ppm cresol with about 80% cresol degradation ability. Immobilized cells showed >99% degradation at high concentration of cresol. The 16S rRNA sequence (accession number: MF278026) deposited in GenBank was used for phylogenetic tree analysis and the strain was grouped under Pseudomonas monteilii. The isolated cresol degrading strain was subsequently named as Pseudomonas monteilii SHY.

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Identification of novel members of the bacterial azoreductase family in Pseudomonas aeruginosa

Cited by 19 publications

References 47 publications

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

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

Azoreductases in drug metabolism

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