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Jean-pierre Jacquot - One of the best experts on this subject based on the ideXlab platform.
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Quinone- and Nitroreductase reactions of Thermotoga maritima peroxiredoxin-Nitroreductase hybrid enzyme.
Archives of Biochemistry and Biophysics, 2012Co-Authors: Zilvinas Anusevicius, Nicolas Rouhier, Jean-pierre Jacquot, Jonas Sarlauskas, Lina Misevičienė, Narimantas ČėnasAbstract:Thermotoga maritima peroxiredoxin–Nitroreductase hybrid enzyme (Prx–NR) consists of a FMN-containing Nitroreductase (NR) domain fused to a peroxiredoxin (Prx) domain. These domains seem to function independently as no electron transfer occurs between them. The reduction of quinones and nitroaromatics by NR proceeded in a two-electron manner, and follows a ‘ping-pong’ scheme with sometimes pronounced inhibition by quinone substrate. The comparison of steady- and presteady-state kinetic data shows that in most cases, the oxidative half-reaction may be rate-limiting in the catalytic cycle of NR. The enzyme was inhibited by dicumarol, a classical inhibitor of oxygen-insensitive Nitroreductases. The reduction of quinones and nitroaromatic compounds by Prx–NR was characterized by the linear dependence of their reactivity (log kcat/Km) on their single-electron reduction potentials E 7^1, while the reactivity of quinones markedly exceeded the one with nitroaromatics. It shows that NR lacks the specificity for the particular structure of these oxidants, except their single-electron accepting potency and the rate of electron self-exchange. It points to the possibility of a single-electron transfer step in a net two-electron reduction of quinones and nitroaromatics by T. maritima Prx–NR, and to a significant diversity of the structures of flavoenzymes which may perform the two-electron reduction of quinones and nitroaromatics
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Quinone- and Nitroreductase reactions of Thermotoga maritima 3 peroxiredoxin–Nitroreductase hybrid enzyme
Archives of Biochemistry and Biophysics, 2012Co-Authors: Zilvinas Anusevicius, Nicolas Rouhier, Lina Misevičienė, Jean-pierre JacquotAbstract:Thermotoga maritima peroxiredoxin–Nitroreductase hybrid enzyme (Prx–NR) consists of a FMN-containing Nitroreductase (NR) domain fused to a peroxiredoxin (Prx) domain. These domains seem to function independently as no electron transfer occurs between them. The reduction of quinones and nitroaromatics by NR proceeded in a two-electron manner, and follows a ‘ping-pong’ scheme with sometimes pronounced inhibition by quinone substrate. The comparison of steady- and presteady-state kinetic data shows that in most cases, the oxidative half-reaction may be rate-limiting in the catalytic cycle of NR. The enzyme was inhibited by dicumarol, a classical inhibitor of oxygen-insensitive Nitroreductases. The reduction of quinones and nitroaromatic compounds by Prx–NR was characterized by the linear dependence of their reactivity (log kcat/Km) on their single-electron reduction potentials E 7^1, while the reactivity of quinones markedly exceeded the one with nitroaromatics. It shows that NR lacks the specificity for the particular structure of these oxidants, except their single-electron accepting potency and the rate of electron self-exchange. It points to the possibility of a single-electron transfer step in a net two-electron reduction of quinones and nitroaromatics by T. maritima Prx–NR, and to a significant diversity of the structures of flavoenzymes which may perform the two-electron reduction of quinones and nitroaromatics
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Two-electron reduction of nitroaromatic compounds by Thermotoga maritima hybrid peroxiredoxin-Nitroreductase enzyme
Chemija, 2011Co-Authors: Nicolas Rouhier, Jean-pierre Jacquot, Lina Miseviciene, Zilvinas Anusevicius, Jonas Sarlauskas, Narimantas CenasAbstract:Explosives such as 2,4,6-trinitrotoluene (TNT) and related polynitroaromatics being toxic environmental pollutants, numerous efforts are directed towards their biodegradation. In this work, we studied the NADPH-dependent two-electron reduction of a number of nitroaromatic compounds by a peroxiredoxin-Nitroreductase hybrid enzyme from Thermotoga maritima (Prx-NR). We have found that the peroxiredoxin and Nitroreductase domains of Prx-NR function independently. The activity of Prx-NR towards nitroaromatics is not influenced by their particular structure and is characterized by a linear log k(cat)/K(m) dependence on their single-electron reduction potentials (E(7)(1)). The reduction of polynitroaromatic explosives N-nitramines such as tetryl (2,4,6-trinitrophenyl-N-methylnitramine) and pentryl (2,4,6-trinitrophenyl-N-nitroaminoethylnitrate) was accompanied by the formation of nitrite, which implies their reductive N-denitration, while p-dinitrobenzene is reduced to p-hydroxylaminonitrobenzene. Taken together, these data indicate that Nitroreductase reactions of Prx-NR share common features with those of other bacterial oxygen-insensitive Nitroreductases, e. g., Enterobacter cloacae Nitroreductase. However, the activity of Prx-NR is 10-100 times lower than that of E. cloacae NR.
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Two-electron reduction of nitroaromatic compounds by Thermotoga maritima hybrid peroxiredoxin- Nitroreductase enzyme
2011Co-Authors: Nicolas Rouhier, Jean-pierre Jacquot, Jonas SarlauskasAbstract:* corresponding author. e-mail: narimantas.cenas@bchi.vu.lt Explosives such as 2,4,6-trinitrotoluene (TNT) and related polynitroaromatics being toxic environmental pollutants, numerous efforts are directed towards their biodegradation. In this work, we studied the NADPH-dependent two-electron reduction of a number of nitroaromatic compounds by a peroxiredoxin–Nitroreductase hybrid enzyme from Thermotoga maritima (Prx–NR). We have found that the peroxiredoxin and Nitroreductase domains of Prx–NR function independently. The activity of Prx–NR towards nitroaromatics is not influenced by their particular structure and is characterized by a linear log kcat / Km dependence on their single-electron reduction potentials (E7). The reduction of polynitroaromatic explosives N-nitramines such as tetryl (2,4,6-trinitrophenyl-N-methylnitramine) and pentryl (2,4,6-trinitrophenyl-N-nitroaminoethylnitrate) was accompanied by the formation of nitrite, which implies their reductive N-denitration, while p-dinitrobenzene is reduced to p-hydroxylaminonitrobenzene. Taken together, these data indicate that Nitroreductase reactions of Prx–NR share common features with those of other bacterial oxygen-insensitive Nitroreductases, e. g., Enterobacter cloacae Nitroreductase. However, the activity of Prx–NR is 10–100 times lower than that of E. cloacae NR.
Takehiko Nohmi - One of the best experts on this subject based on the ideXlab platform.
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Purification and Characterization of Wild-type and Mutant “Classical” Nitroreductases of Salmonella typhimurium L33R MUTATION GREATLY DIMINISHES BINDING OF FMN TO THE Nitroreductase OF S. TYPHIMURIUM
Journal of Biological Chemistry, 1998Co-Authors: Masahiko Watanabe, Tatsuya Nishino, Koji Takio, Toshio Sofuni, Takehiko NohmiAbstract:Abstract “Classical” Nitroreductase ofSalmonella typhimurium is a flavoprotein that catalyzes the reduction of nitroaromatics to metabolites that are toxic, mutagenic, or carcinogenic. This enzyme represents a new class of flavin-dependent enzymes, which includes Nitroreductases ofEnterobacter cloacae and Escherichia coli,flavin oxidoreductase of Vibrio fischeri, and NADH oxidase of Thermus thermophilus. To investigate the structure-function relation of this class of enzymes, the gene encoding a mutant Nitroreductase was cloned from S. typhimuriumstrain TA1538NR, and the enzymatic properties were compared with those of the wild-type. DNA sequence analysis revealed a T to G mutation in the mutant Nitroreductase gene, predicting a replacement of leucine 33 with arginine. In contrast to the wild-type enzyme, the purified protein with a mutation of leucine 33 to arginine has no detectable Nitroreductase activities in the standard assay conditions and easily lost FMN by dialysis or ultrafiltration. In the presence of an excess amount of FMN, however, the mutant protein exhibited a weak but measurable enzyme activity, and the substrate specificity was similar to that of the wild-type enzyme. Possible mechanisms by which the mutation greatly diminishes binding of FMN to the Nitroreductase are discussed.
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Purification and Characterization of Wild-type and Mutant "Classical" Nitroreductases of Salmonella typhimurium
1998Co-Authors: Masahiko Watanabe, Tatsuya Nishino, Toshio Sofuni, Koji Takioi, Takehiko NohmiAbstract:“Classical” Nitroreductase of Salmonella typhimurium is a flavoprotein that catalyzes the reduction of nitroaromatics to metabolites that are toxic, mutagenic, or carcinogenic. This enzyme represents a new class of flavin-dependent enzymes, which includes Nitroreductases of Enterobacter cloacae and Escherichia coli, flavin oxidoreductase of Vibrio fischeri, and NADH oxidase of Thermus thermophilus. To investigate the structurefunction relation of this class of enzymes, the gene encoding a mutant Nitroreductase was cloned from S. typhimurium strain TA1538NR, and the enzymatic properties were compared with those of the wild-type. DNA sequence analysis reveale daTt o Gmutation in the mutant Nitroreductase gene, predicting a replacement of leucine 33 with arginine. In contrast to the wild-type enzyme, the purified protein with a mutation of leucine 33 to arginine has no detectable Nitroreductase activities in the standard assay conditions and easily lost FMN by dialysis or ultrafiltration. In the presence of an excess amount of FMN, however, the mutant protein exhibited a weak but measurable enzyme activity, and the substrate specificity was similar to that of the wildtype enzyme. Possible mechanisms by which the mutation greatly diminishes binding of FMN to the Nitroreductase are discussed.
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targeted disruption of the gene encoding the classical Nitroreductase enzyme in salmonella typhimurium ames test strains ta1535 and ta1538
Mutation Research, 1997Co-Authors: Masami Yamada, Masahiko Watanabe, Toshio Sofuni, Javier Espinosaaguirre, Keiko Matsui, Takehiko NohmiAbstract:The gene encoding the 'classical Nitroreductase' (CNR) of Salmonella typhimurium was disrupted. In this manner, cnr null mutant derivatives of strains TA1535 and TA1538 were constructed, and named YG7131 and YG7127, respectively. In both strain backgrounds, cnr gene disruption reduced nitrofurazone-reductase activity. This reduction almost completely eliminated the Nitroreductase activity of strain TA1538. In contrast, the Nitroreductase activity of strain TA1535 was much higher than that in TA1538. In this background, cnr gene disruption resulted in a reduction in Nitroreductase activity by a similar absolute amount as in TA1538, but representing only about one-quarter of the original activity of TA1535. The results suggest that S. typhimurium has originally at least two distinct Nitroreductases, one of which is already deficient in strain TA1538; the CNR is present in both TA1535 and TA1538. Also, these two strains (including their derivatives, TA98 and TA100) are not isogenic with regard to Nitroreductase activity. After the introduction of plasmid pKM101, the sensitivities of the strains YG7132 and YG7128, the cnr-null mutants of TA98 and TA100, respectively, against several nitro compounds were compared with those of the conventional cnr-deficient strains TA98NR and TA100NR and the wild-type strains TA98 and TA100. The mutagenicities of 2-nitrofluorene and 1-nitropyrene in YG7132 or TA98NR were ten-fold lower than those of the compounds in TA98. Similarly, the mutagenicity of 2-(2-furyl)-3-(5-nitro-2-furyl) acrylamide in strain YG7128 or TA100NR was substantially lower than that of the compound in TA100. However, the mutagenicity of 2-nitronaphthalene in YG7128 was between those observed with TA100 and TA100NR, suggesting that a Nitroreductase in S. typhimurium other than CNR is involved in the metabolic activation of this compound. The cnr gene of S. typhimurium positively hybridized with DNA at 13 min on the E. coli chromosome where the nfsB and nfnB genes of E. coli are mapped. These results suggest that the cnr gene of S. typhimurium is a counterpart of the nfsB and nfnB genes of E. coli, and that the newly constructed cnr-deletion strains are useful to assess the role of Nitroreductases in the metabolic activation of mutagenic nitro compounds.
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Nucleotide sequence of Salmonella typhimurium Nitroreductase gene
Nucleic Acids Research, 1990Co-Authors: Masahiko Watanabe, Motoi Ishidate, Takehiko NohmiAbstract:'Classical Nitroreductase' is an enzyme involved in theintracellular metabolic activation ofmutagenic nitroarenes (1).Wehave already cloned the Nitroreductase gene ofSalmonellatyphimuriumTA1538(2) anddeterminedthenucleotide sequenceby dideoxy sequence analysis ofboth strands. Below is shownthe nucleotide sequence of 1690base fragment which containsanopenreadingframeof651 nucleotides withpotential to encodethe Nitroreductase. Themaxicell technique wasusedto identifythe Nitroreductase and its molecular weight was estimated as28KDa, which is close to the calculated molecular weight of23,955. Possible sequence of -35, -10, S.D. and rho-independent transcriptional termination signal are indicated.
Hong-gyu Song - One of the best experts on this subject based on the ideXlab platform.
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Nitroreductase II involved in 2,4,6-trinitrotoluene degradation: purification and characterization from Klebsiella sp. Cl.
The Journal of Microbiology, 2009Co-Authors: Jung-hye Shin, Hong-gyu SongAbstract:Three 2,4,6-trinitrotoluene (TNT) Nitroreductases from Klebsiella sp. CI have different reduction capabilities that can degrade TNT by simultaneous utilization of two initial reduction pathways. Of these, Nitroreductase II was purified to homogeneity by sequential chromatographies. Nitroreductase II is an oxygen-insensitive enzyme and reduces both TNT and nitroblue tetrazolium. The N-terminal amino acid sequence of the enzyme did not show any sequence similarity with those of other Nitroreductases reported. However, it transformed TNT by the reduction of nitro groups like Nitroreductase I. It had a higher substrate affinity and specific activity for TNT reduction than other Nitroreductases, and it showed a higher oxidation rate of NADPH with the ortho-substituted isomers of TNT metabolites (2-hydroxylaminodinitrotoluene and 2-aminodinitrotoluene) than with para-substituted compounds (4-hydroxylaminodinitrotoluene and 4-amino-dinitrotoluene).
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Purification and characterization of NAD(P)H-dependent Nitroreductase I from Klebsiella sp. C1 and enzymatic transformation of 2,4,6-trinitrotoluene
Applied Microbiology and Biotechnology, 2005Co-Authors: Hong-gyu SongAbstract:Three NAD(P)H-dependent Nitroreductases that can transform 2,4,6-trinitrotoluene (TNT) by two reduction pathways were detected in Klebsiella sp. C1. Among these enzymes, the protein with the highest reduction activity of TNT (Nitroreductase I) was purified to homogeneity using ion-exchange, hydrophobic interaction, and size exclusion chromatographies. Nitroreductase I has a molecular mass of 27 kDa as determined by SDS-PAGE, and exhibits a broad pH optimum between 5.5 and 6.5, with a temperature optimum of 30–40°C. Flavin mononucleotide is most likely the natural flavin cofactor of this enzyme. The N-terminal amino acid sequence of this enzyme does not show a high degree of sequence similarity with Nitroreductases from other enteric bacteria. This enzyme catalyzed the two-electron reduction of several nitroaromatic compounds with very high specific activities of NADPH oxidation. In the enzymatic transformation of TNT, 2-amino-4,6-dinitrotoluene and 2,2′,6,6′-tetranitro-4,4′-azoxytoluene were detected as transformation products. Although this bacterium utilizes the direct ring reduction and subsequent denitration pathway together with a nitro group reduction pathway, metabolites in direct ring reduction of TNT could not easily be detected. Unlike other Nitroreductases, Nitroreductase I was able to transform hydroxylaminodinitrotoluenes (HADNT) into aminodinitrotoluenes (ADNT), and could reduce ortho isomers (2-HADNT and 2-ADNT) more easily than their para isomers (4-HADNT and 4-ADNT). Only the nitro group in the ortho position of 2,4-DNT was reduced to produce 2-hydroxylamino-4-nitrotoluene by Nitroreductase I; the nitro group in the para position was not reduced.
Wen Shi - One of the best experts on this subject based on the ideXlab platform.
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A Lysosome-Targeting Fluorescence Off-On Probe for Imaging of Nitroreductase and Hypoxia in Live Cells.
Chemistry - An Asian Journal, 2016Co-Authors: Jin Zhou, Wen Shi, Qiuyu GongAbstract:A lysosome-targeting fluorescent off-on probe has been developed by one-step synthesis for detecting lysosomal Nitroreductase and hypoxia. The probe is constructed by incorporating morpholine (a lysosome-targeting unit) into 4-nitro-1,8-naphthalimide (as a fluorochrome and specific substrate for Nitroreductase), and the detection mechanism is based on the Nitroreductase-catalyzed reduction of the probe to 4-amino-1,8-naphthalimide, accompanied by a large fluorescence enhancement at a wavelength of 543 nm. The probe shows an accurate lysosome-targeting ability with high selectivity and sensitivity to Nitroreductase (detection limit: 2.2 ng mL−1). Notably, the probe has been used to image the change of lysosomal Nitroreductase in live cells during hypoxia, revealing that the increase of Nitroreductase in lysosomes may be smaller than that in the cytoplasm. In addition, the probe is expected to be useful for studying the function of Nitroreductase in the acidic organelle of lysosomes.
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in vivo imaging and detection of Nitroreductase in zebrafish by a new near infrared fluorescence off on probe
Biosensors and Bioelectronics, 2015Co-Authors: Zhe Wang, Ronghua Yang, Wen ShiAbstract:Abstract A new near-infrared fluorescence off–on probe is developed and applied to fluorescence imaging of Nitroreductase in zebrafish in vivo . The probe is readily prepared by connecting 4-nitrobenzene as a quenching and recognizing moiety to a stable hemicyanine skeleton that can be formed via the decomposition of IR 780. The fluorescence off–on response of the probe to Nitroreductase is based on the enzyme-catalyzed reduction of the 4-nitrobenzene moiety, followed by the 1,6-rearrangement-elimination and the fluorophore release. Compared with the existing Nitroreductase probes, the proposed probe exhibits superior analytical performance such as near-infrared fluorescence emission over 700 nm as well as high selectivity and sensitivity, with a detection limit of 14 ng/mL. More importantly, the probe has been successfully applied to visualize the distribution of Nitroreductase in living zebrafish in vivo , revealing that Nitroreductase might mainly exist in zebrafish yolk sac. The superior properties of the probe make it of great potential use in other biosystems and in vivo studies.
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in vivo imaging and detection of Nitroreductase in zebrafish by a new near-infrared fluorescence off–on probe
Biosensors and Bioelectronics, 2015Co-Authors: Zhe Wang, Ronghua Yang, Wen ShiAbstract:Abstract A new near-infrared fluorescence off–on probe is developed and applied to fluorescence imaging of Nitroreductase in zebrafish in vivo . The probe is readily prepared by connecting 4-nitrobenzene as a quenching and recognizing moiety to a stable hemicyanine skeleton that can be formed via the decomposition of IR 780. The fluorescence off–on response of the probe to Nitroreductase is based on the enzyme-catalyzed reduction of the 4-nitrobenzene moiety, followed by the 1,6-rearrangement-elimination and the fluorophore release. Compared with the existing Nitroreductase probes, the proposed probe exhibits superior analytical performance such as near-infrared fluorescence emission over 700 nm as well as high selectivity and sensitivity, with a detection limit of 14 ng/mL. More importantly, the probe has been successfully applied to visualize the distribution of Nitroreductase in living zebrafish in vivo , revealing that Nitroreductase might mainly exist in zebrafish yolk sac. The superior properties of the probe make it of great potential use in other biosystems and in vivo studies.
Nicolas Rouhier - One of the best experts on this subject based on the ideXlab platform.
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Quinone- and Nitroreductase reactions of Thermotoga maritima peroxiredoxin-Nitroreductase hybrid enzyme.
Archives of Biochemistry and Biophysics, 2012Co-Authors: Zilvinas Anusevicius, Nicolas Rouhier, Jean-pierre Jacquot, Jonas Sarlauskas, Lina Misevičienė, Narimantas ČėnasAbstract:Thermotoga maritima peroxiredoxin–Nitroreductase hybrid enzyme (Prx–NR) consists of a FMN-containing Nitroreductase (NR) domain fused to a peroxiredoxin (Prx) domain. These domains seem to function independently as no electron transfer occurs between them. The reduction of quinones and nitroaromatics by NR proceeded in a two-electron manner, and follows a ‘ping-pong’ scheme with sometimes pronounced inhibition by quinone substrate. The comparison of steady- and presteady-state kinetic data shows that in most cases, the oxidative half-reaction may be rate-limiting in the catalytic cycle of NR. The enzyme was inhibited by dicumarol, a classical inhibitor of oxygen-insensitive Nitroreductases. The reduction of quinones and nitroaromatic compounds by Prx–NR was characterized by the linear dependence of their reactivity (log kcat/Km) on their single-electron reduction potentials E 7^1, while the reactivity of quinones markedly exceeded the one with nitroaromatics. It shows that NR lacks the specificity for the particular structure of these oxidants, except their single-electron accepting potency and the rate of electron self-exchange. It points to the possibility of a single-electron transfer step in a net two-electron reduction of quinones and nitroaromatics by T. maritima Prx–NR, and to a significant diversity of the structures of flavoenzymes which may perform the two-electron reduction of quinones and nitroaromatics
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Quinone- and Nitroreductase reactions of Thermotoga maritima 3 peroxiredoxin–Nitroreductase hybrid enzyme
Archives of Biochemistry and Biophysics, 2012Co-Authors: Zilvinas Anusevicius, Nicolas Rouhier, Lina Misevičienė, Jean-pierre JacquotAbstract:Thermotoga maritima peroxiredoxin–Nitroreductase hybrid enzyme (Prx–NR) consists of a FMN-containing Nitroreductase (NR) domain fused to a peroxiredoxin (Prx) domain. These domains seem to function independently as no electron transfer occurs between them. The reduction of quinones and nitroaromatics by NR proceeded in a two-electron manner, and follows a ‘ping-pong’ scheme with sometimes pronounced inhibition by quinone substrate. The comparison of steady- and presteady-state kinetic data shows that in most cases, the oxidative half-reaction may be rate-limiting in the catalytic cycle of NR. The enzyme was inhibited by dicumarol, a classical inhibitor of oxygen-insensitive Nitroreductases. The reduction of quinones and nitroaromatic compounds by Prx–NR was characterized by the linear dependence of their reactivity (log kcat/Km) on their single-electron reduction potentials E 7^1, while the reactivity of quinones markedly exceeded the one with nitroaromatics. It shows that NR lacks the specificity for the particular structure of these oxidants, except their single-electron accepting potency and the rate of electron self-exchange. It points to the possibility of a single-electron transfer step in a net two-electron reduction of quinones and nitroaromatics by T. maritima Prx–NR, and to a significant diversity of the structures of flavoenzymes which may perform the two-electron reduction of quinones and nitroaromatics
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Two-electron reduction of nitroaromatic compounds by Thermotoga maritima hybrid peroxiredoxin-Nitroreductase enzyme
Chemija, 2011Co-Authors: Nicolas Rouhier, Jean-pierre Jacquot, Lina Miseviciene, Zilvinas Anusevicius, Jonas Sarlauskas, Narimantas CenasAbstract:Explosives such as 2,4,6-trinitrotoluene (TNT) and related polynitroaromatics being toxic environmental pollutants, numerous efforts are directed towards their biodegradation. In this work, we studied the NADPH-dependent two-electron reduction of a number of nitroaromatic compounds by a peroxiredoxin-Nitroreductase hybrid enzyme from Thermotoga maritima (Prx-NR). We have found that the peroxiredoxin and Nitroreductase domains of Prx-NR function independently. The activity of Prx-NR towards nitroaromatics is not influenced by their particular structure and is characterized by a linear log k(cat)/K(m) dependence on their single-electron reduction potentials (E(7)(1)). The reduction of polynitroaromatic explosives N-nitramines such as tetryl (2,4,6-trinitrophenyl-N-methylnitramine) and pentryl (2,4,6-trinitrophenyl-N-nitroaminoethylnitrate) was accompanied by the formation of nitrite, which implies their reductive N-denitration, while p-dinitrobenzene is reduced to p-hydroxylaminonitrobenzene. Taken together, these data indicate that Nitroreductase reactions of Prx-NR share common features with those of other bacterial oxygen-insensitive Nitroreductases, e. g., Enterobacter cloacae Nitroreductase. However, the activity of Prx-NR is 10-100 times lower than that of E. cloacae NR.
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Two-electron reduction of nitroaromatic compounds by Thermotoga maritima hybrid peroxiredoxin- Nitroreductase enzyme
2011Co-Authors: Nicolas Rouhier, Jean-pierre Jacquot, Jonas SarlauskasAbstract:* corresponding author. e-mail: narimantas.cenas@bchi.vu.lt Explosives such as 2,4,6-trinitrotoluene (TNT) and related polynitroaromatics being toxic environmental pollutants, numerous efforts are directed towards their biodegradation. In this work, we studied the NADPH-dependent two-electron reduction of a number of nitroaromatic compounds by a peroxiredoxin–Nitroreductase hybrid enzyme from Thermotoga maritima (Prx–NR). We have found that the peroxiredoxin and Nitroreductase domains of Prx–NR function independently. The activity of Prx–NR towards nitroaromatics is not influenced by their particular structure and is characterized by a linear log kcat / Km dependence on their single-electron reduction potentials (E7). The reduction of polynitroaromatic explosives N-nitramines such as tetryl (2,4,6-trinitrophenyl-N-methylnitramine) and pentryl (2,4,6-trinitrophenyl-N-nitroaminoethylnitrate) was accompanied by the formation of nitrite, which implies their reductive N-denitration, while p-dinitrobenzene is reduced to p-hydroxylaminonitrobenzene. Taken together, these data indicate that Nitroreductase reactions of Prx–NR share common features with those of other bacterial oxygen-insensitive Nitroreductases, e. g., Enterobacter cloacae Nitroreductase. However, the activity of Prx–NR is 10–100 times lower than that of E. cloacae NR.
