Search Results (16)

In order to detail the antiplasmodial effects of quinones (Q) and nitroaromatic compounds (ArNO₂), we investigated their reduction mechanism by Plasmodium falciparum flavoenzyme type II NADH:ubiquinone oxidoreductase (PfNDH2). The reactivity of Q and ArNO₂ (n = 29) follows a common trend and exhibits a parabolic dependence on their single-electron reduction potential ($E_7^1$), albeit with significantly scattered data. The reactivity of quinones with similar $E_7^1$ values increases with their lipophilicity. Quinones are reduced by PfNDH2 in a two-electron way, but ArNO₂ are reduced in a single-electron way. The inhibition studies using NAD⁺ and ADP-ribose showed that quinones oxidize the complexes of reduced enzyme with NADH and NAD⁺. This suggests that, as in the case of other NDH2s, quinones and the nicotinamide ring of NAD(H) bind at separate sites. A scheme of PfNDH2 catalysis is proposed, consistent with both the observed ‘ping-pong’ mechanism and the presence of two substrate binding sites. Molecular docking showed that Q and ArNO₂ bind in a similar manner and that lipophilic quinones have a higher affinity for the binding site. One may expect that PfNDH2 can be partially responsible for the previously observed enhanced antiplasmodial activity of aziridinylbenzoquinones caused by their two-electron reduction, as well as for the redox cycling and oxidative stress-type action of ArNO₂. Full article

Bacillus subtilis ferredoxin:NADP⁺ oxidoreductase (BsFNR) is a thioredoxin reductase-type FNR whose redox properties and reactivity with nonphysiological electron acceptors have been scarcely characterized. On the basis of redox reactions with 3-acetylpyridine adenine dinucleotide phosphate, the two-electron reduction midpoint potential of the flavin adenine dinucleotide (FAD) cofactor was estimated to be −0.240 V. Photoreduction using 5-deazaflavin mononucleotide (5-deazaFMN) as a photosensitizer revealed that the difference in the redox potentials between the first and second single-electron transfer steps was 0.024 V. We examined the mechanisms of the reduction of several different groups of non-physiological electron acceptors catalyzed by BsFNR. The reactivity of quinones and aromatic N-oxides toward BsFNR increased when increasing their single-electron reduction midpoint redox potentials. The reactivity of nitroaromatic compounds was lower due to their lower electron self-exchange rate, but it exhibited the same trend. A mixed single- and two-electron reduction reaction was characteristic of quinones, whereas reactions involving nitroaromatics proceeded exclusively via the one-electron reduction reaction. The oxidation of FADH^• to FAD is the rate-limiting step during the oxidation of fully reduced FAD. The calculated electron transfer distances in the reaction with nitroaromatics were close to those of other FNRs including the plant-type enzymes, thus demonstrating their similar active site accessibility to low-molecular-weight oxidants despite the fundamental differences in their structures. Full article

E. coli nitroreductase A (NfsA) is a candidate for gene-directed prodrug cancer therapy using bioreductively activated nitroaromatic compounds (ArNO₂). In this work, we determined the standard redox potential of FMN of NfsA to be −215 ± 5 mV at pH 7.0. FMN semiquinone was not formed during 5-deazaflavin-sensitized NfsA photoreduction. This determines the two-electron character of the reduction of ArNO₂ and quinones (Q). In parallel, we characterized the oxidant specificity of NfsA with an emphasis on its structure. Except for negative outliers nitracrine and SN-36506, the reactivity of ArNO₂ increases with their electron affinity (single-electron reduction potential, E¹₇) and is unaffected by their lipophilicity and Van der Waals volume up to 386 Å. The reactivity of quinoidal oxidants is not clearly dependent on E¹₇, but 2-hydroxy-1,4-naphthoquinones were identified as positive outliers and a number of compounds with diverse structures as negative outliers. 2-Hydroxy-1,4-naphthoquinones are characterized by the most positive reaction activation entropy and the negative outlier tetramethyl-1,4-benzoquinone by the most negative. Computer modelling data showed that the formation of H bonds with Arg15, Arg133, and Ser40, plays a major role in the binding of oxidants to reduced NfsA, while the role of the π–π interaction of their aromatic structures is less significant. Typically, the calculated hydride-transfer distances during ArNO₂ reduction are smallwer than for Q. This explains the lower reactivity of quinones. Another factor that slows down the reduction is the presence of positively charged aliphatic substituents. Full article

Xenobiotic reductase B (XenB) catalyzes the reduction of the aromatic ring or nitro groups of nitroaromatic compounds with methyl, amino or hydroxyl radicals. This reaction is of biotechnological interest for bioremediation, the reuse of industrial waste or the activation of prodrugs. However, the structural factors that explain the binding of XenB to different substrates are unknown. Molecular dynamics simulations and quantum mechanical calculations were performed to identify the residues involved in the formation and stabilization of the enzyme/substrate complex and to explain the use of different substrates by this enzyme. Our results show that Tyr65 and Tyr335 residues stabilize the ligands through hydrophobic interactions mediated by the aromatic rings of these aminoacids. The higher XenB activity determined with the substrates 1,3,5-trinitrobenzene and 2,4,6-trinitrotoluene is consistent with the lower energy of the highest occupied molecular orbital (LUMO) orbitals and a lower energy of the homo orbital (LUMO), which favors electrophile and nucleophilic activity, respectively. The electrostatic potential maps of these compounds suggest that the bonding requires a large hydrophobic region in the aromatic ring, which is promoted by substituents in ortho and para positions. These results are consistent with experimental data and could be used to propose point mutations that allow this enzyme to process new molecules of biotechnological interest. Full article

Doping of ferrites is an important domain of research for their application as photocatalysts. In the present work, the effect of Ti⁴⁺ substitution on the structural and photocatalytic properties of strontium ferrite nanoparticles (NPs) is studied. Ternary doped Sr_1−xTi_xFe₂O_4+δ ferrite NPs (x = 0.0–1.0) were synthesized by sol–gel methodology. Tetravalent Ti⁴⁺ ions caused oxygen hyperstoichiometry and enhancement in the surface area from 44.3 m²/g for SrFe₂O₄ NPs to 77.6 m²/g for Sr_0.4Ti_0.6Fe₂O_4+δ NPs. The average diameter of NPs ranged between 25–35 nm as revealed by TEM analysis. The presence of two sextets in the Mössbauer spectrum of pristine SrFe₂O₄ and Ti⁴⁺-substituted ferrite NPs and a paramagnetic doublet in the TiFe₂O₅ confirmed their phase purity. The photocatalytic potential of pure and Ti⁴⁺-substituted ferrite NPs was studied using nitroaromatic compounds, viz. pendimethalin, p-nitrophenol and Martius yellow, as model pollutants. Doped ferrite NPs with a composition of Sr_0.4Ti_0.6Fe₂O_4+δ NPs showed the highest degradation efficiency ranging from 87.2% to 94.4%. The increased photocatalytic potential was ascribed to the lowering of band gap (E_g) from 2.45 eV to 2.18 eV, a fourfold decrease in photoluminescence intensity, increased charge carrier concentration (4.90 × 10¹⁵ cm⁻³ to 6.96 × 10¹⁵ cm⁻³), and decreased barrier height from 1.20 to 1.02 eV. O₂^●− radicals appeared to be the main reactive oxygen species involved in photodegradation. The apparent rate constant values using the Langmuir–Hinshelwood kinetic model were 1.9 × 10⁻² min⁻¹, 2.3 × 10⁻² min⁻¹ and 1.3 × 10⁻² min⁻¹ for p-nitrophenol, pendimethalin and Martius yellow, respectively. Thus, tuning the Ti⁴⁺ content in strontium ferrite NPs proved to be an effective strategy in improving their photocatalytic potential for the degradation of nitroaromatic pollutants. Full article

Rhodopseudomonas palustris ferredoxin:NADP⁺ oxidoreductase (RpFNR) belongs to a novel group of thioredoxin reductase-type FNRs with partly characterized redox properties. Based on the reactions of RpFNR with the 3-acetylpyridine adenine dinucleotide phosphate redox couple, we estimated the two-electron reduction midpoint potential of the FAD cofactor to be −0.285 V. 5-Deaza-FMN-sensitized photoreduction revealed −0.017 V separation of the redox potentials between the first and second electron transfer events. We examined the mechanism of oxidation of RpFNR by several different groups of nonphysiological electron acceptors. The k_cat/K_m values of quinones and aromatic N-oxides toward RpFNR increase with their single-electron reduction midpoint potential. The lower reactivity, mirroring their lower electron self-exchange rate, is also seen to have a similar trend for nitroaromatic compounds. A mixed single- and two-electron reduction was characteristic of quinones, with single-electron reduction accounting for 54% of the electron flux, whereas nitroaromatics were reduced exclusively via single-electron reduction. It is highly possible that the FADH· to FAD oxidation reaction is the rate-limiting step during the reoxidation of reduced FAD. The calculated electron transfer distances in the reaction with quinones and nitroaromatics were close to those of Anabaena and Plasmodium falciparum FNRs, thus demonstrating their similar “intrinsic” reactivity. Full article

Neuronal nitric oxide synthase (nNOS) catalyzes single-electron reduction of quinones (Q), nitroaromatic compounds (ArNO₂) and aromatic N-oxides (ArN → O), and is partly responsible for their oxidative stress-type cytotoxicity. In order to expand a limited knowledge on the enzymatic mechanisms of these processes, we aimed to disclose the specific features of nNOS in the reduction of such xenobiotics. In the absence or presence of calmodulin (CAM), the reactivity of Q and ArN → O increases with their single-electron reduction midpoint potential (E¹₇). ArNO₂ form a series with lower reactivity. The calculations according to an “outer-sphere” electron transfer model show that the binding of CAM decreases the electron transfer distance from FMNH₂ to quinone by 1–2 Å. The effects of ionic strength point to the interaction of oxidants with a negatively charged protein domain close to FMN, and to an increase in accessibility of the active center induced by high ionic strength. The multiple turnover experiments of nNOS show that, in parallel with reduced FAD-FMN, duroquinone reoxidizes the reduced heme, in particular its Fe²⁺-NO form. This finding may help to design the heme-targeted bioreductively activated agents and contribute to the understanding of the role of P-450-type heme proteins in the bioreduction of quinones and other prooxidant xenobiotics. Full article

In this paper, substituted anilines are industrially obtained by direct hydrogenation of nitroaromatic compounds with molecular H₂ using metals as catalysts. Previous theoretical studies proposed that the mechanism of the reaction depends on the nature of the metal used as a catalyst, and that rationally designed bimetallic materials might show improved catalytic performance. Herein, we present IR spectroscopic studies of nitrobenzene interactions with monometallic Ni/SiO₂, Cu/SiO₂ and Pd/SiO_2, and with bimetallic CuNi/SiO₂ and CuPd/SiO₂ catalysts, both in the absence and presence of H₂, combined with density functional theory (DFT) calculations on selected bimetallic NiCu(111) and PdCu(111) models. The results obtained experimentally confirm that the reaction mechanism on non-noble metals such as Ni proceeds through N-O bond dissociation, generating nitrosobenzene intermediates, while, on noble metals, such as Pd, H-attack is necessary to activate the NO bond. Moreover, a bimetallic CuPd/SiO₂ catalyst with a Pd enriched surface is prepared that exhibits an enhanced H₂ dissociation ability and a particular reactivity at the boundary between the two metals. Full article

Nitroaromatic compounds (ArNO₂) maintain their importance in relation to industrial processes, environmental pollution, and pharmaceutical application. The manifestation of toxicity/therapeutic action of nitroaromatics may involve their single- or two-electron reduction performed by various flavoenzymes and/or their physiological redox partners, metalloproteins. The pivotal and still incompletely resolved questions in this area are the identification and characterization of the specific enzymes that are involved in the bioreduction of ArNO₂ and the establishment of their contribution to cytotoxic/therapeutic action of nitroaromatics. This review addresses the following topics: (i) the intrinsic redox properties of ArNO₂, in particular, the energetics of their single- and two-electron reduction in aqueous medium; (ii) the mechanisms and structure-activity relationships of reduction in ArNO₂ by flavoenzymes of different groups, dehydrogenases-electrontransferases (NADPH:cytochrome P-450 reductase, ferredoxin:NADP(H) oxidoreductase and their analogs), mammalian NAD(P)H:quinone oxidoreductase, bacterial nitroreductases, and disulfide reductases of different origin (glutathione, trypanothione, and thioredoxin reductases, lipoamide dehydrogenase), and (iii) the relationships between the enzymatic reactivity of compounds and their activity in mammalian cells, bacteria, and parasites. Full article

Nitroaromatic antibiotics show activity against anaerobic bacteria and parasites, finding use in the treatment of Heliobacter pylori infections, tuberculosis, trichomoniasis, human African trypanosomiasis, Chagas disease and leishmaniasis. Despite this activity and a clear need for the development of new treatments for these conditions, the associated toxicity and lack of clear mechanisms of action have limited their therapeutic development. Nitroaromatic antibiotics require reductive bioactivation for activity and this reductive metabolism can convert the nitro group to nitric oxide (NO) or a related reactive nitrogen species (RNS). As nitric oxide plays important roles in the defensive immune response to bacterial infection through both signaling and redox-mediated pathways, defining controlled NO generation pathways from these antibiotics would allow the design of new therapeutics. This review focuses on the release of nitrogen oxide species from various nitroaromatic antibiotics to portend the increased ability for these compounds to positively impact infectious disease treatment. Full article

Ferredoxin:NADP⁺ oxidoreductase from Plasmodium falciparum (PfFNR) catalyzes the NADPH-dependent reduction of ferredoxin (PfFd), which provides redox equivalents for the biosynthesis of isoprenoids and fatty acids in the apicoplast. Like other flavin-dependent electrontransferases, PfFNR is a potential source of free radicals of quinones and other redox cycling compounds. We report here a kinetic study of the reduction of quinones, nitroaromatic compounds and aromatic N-oxides by PfFNR. We show that all these groups of compounds are reduced in a single-electron pathway, their reactivity increasing with the increase in their single-electron reduction midpoint potential (E¹₇). The reactivity of nitroaromatics is lower than that of quinones and aromatic N-oxides, which is in line with the differences in their electron self-exchange rate constants. Quinone reduction proceeds via a ping-pong mechanism. During the reoxidation of reduced FAD by quinones, the oxidation of FADH^. to FAD is the possible rate-limiting step. The calculated electron transfer distances in the reaction of PfFNR with various electron acceptors are similar to those of Anabaena FNR, thus demonstrating their similar “intrinsic” reactivity. Ferredoxin stimulated quinone- and nitro-reductase reactions of PfFNR, evidently providing an additional reduction pathway via reduced PfFd. Based on the available data, PfFNR and possibly PfFd may play a central role in the reductive activation of quinones, nitroaromatics and aromatic N-oxides in P. falciparum, contributing to their antiplasmodial action. Full article

With the aim to clarify the mechanism(s) of action of nitroaromatic compounds against the malaria parasite Plasmodium falciparum, we examined the single-electron reduction by P. falciparum ferredoxin:NADP⁺ oxidoreductase (PfFNR) of a series of nitrofurans and nitrobenzenes (n = 23), and their ability to inhibit P. falciparum glutathione reductase (PfGR). The reactivity of nitroaromatics in PfFNR-catalyzed reactions increased with their single-electron reduction midpoint potential (E¹₇). Nitroaromatic compounds acted as non- or uncompetitive inhibitors towards PfGR with respect to NADPH and glutathione substrates. Using multiparameter regression analysis, we found that the in vitro activity of these compounds against P. falciparum strain FcB1 increased with their E¹₇ values, octanol/water distribution coefficients at pH 7.0 (log D), and their activity as PfGR inhibitors. Our data demonstrate that both factors, the ease of reductive activation and the inhibition of PfGR, are important in the antiplasmodial in vitro activity of nitroaromatics. To the best of our knowledge, this is the first quantitative demonstration of this kind of relationship. No correlation between antiplasmodial activity and ability to inhibit human erythrocyte GR was detected in tested nitroaromatics. Our data suggest that the efficacy of prooxidant antiparasitic agents may be achieved through their combined action, namely inhibition of antioxidant NADPH:disulfide reductases, and the rapid reduction by single-electron transferring dehydrogenases-electrontransferases. Full article

Nitroaromatic compounds—adducts of Morita–Baylis–Hillman (MBHA) reaction—have been applied in the treatment of malaria, leishmaniasis, and Chagas disease. The biological activity of these compounds is directly related to chemical reactivity in the environment, chemical structure of the compound, and reduction of the nitro group. Because of the last aspect, electrochemical methods are used to simulate the pharmacological activity of nitroaromatic compounds. In particular, previous studies have shown a correlation between the one-electron reduction potentials in aprotic medium (estimated by cyclic voltammetry) and antileishmanial activities (measured by the IC₅₀) for a series of twelve MBHA. In the present work, two different computational protocols were calibrated to simulate the reduction potentials for this series of molecules with the aim of supporting the molecular modeling of new pharmacological compounds from the prediction of their reduction potentials. The results showed that it was possible to predict the experimental reduction potential for the calibration set with mean absolute errors of less than 25 mV (about 0.6 kcal·mol⁻¹). Full article

Nitroreductases (NRs) hold promise for converting nitroaromatics to aromatic amines. Nitroaromatic reduction rate increases with Hammett substituent constant for NRs from two different subgroups, confirming substrate identity as a key determinant of reactivity. Amine yields were low, but compounds yielding amines tend to have a large ? system and electron withdrawing substituents. Therefore, we also assessed the prospects of varying the enzyme. Several different subgroups of NRs include members able to produce aromatic amines. Comparison of four NR subgroups shows that they provide contrasting substrate binding cavities with distinct constraints on substrate position relative to the flavin. The unique architecture of the NR dimer produces an enormous contact area which we propose provides the stabilization needed to offset the costs of insertion of the active sites between the monomers. Thus, we propose that the functional diversity included in the NR superfamily stems from the chemical versatility of the flavin cofactor in conjunction with a structure that permits tremendous active site variability. These complementary properties make NRs exceptionally promising enzymes for development for biocatalysis in prodrug activation and conversion of nitroaromatics to valuable aromatic amines. We provide a framework for identifying NRs and substrates with the greatest potential to advance. Full article

The enzymatic reactivity of a series of benzo[1,2-c]1,2,5-oxadiazole N-oxides (benzofuroxans; BFXs) towards mammalian single-electron transferring NADPH:cytochrome P-450 reductase (P-450R) and two-electron (hydride) transferring NAD(P)H:quinone oxidoreductase (NQO1) was examined in this work. Since the =N⁺ (?O)O^? moiety of furoxan fragments of BFXs bears some similarity to the aromatic nitro-group, the reactivity of BFXs was compared to that of nitro-aromatic compounds (NACs) whose reduction mechanisms by these and other related flavoenzymes have been extensively investigated. The reduction of BFXs by both P-450R and NQO1 was accompanied by O₂ uptake, which was much lower than the NADPH oxidation rate; except for annelated BFXs, whose reduction was followed by the production of peroxide. In order to analyze the possible quantitative structure-activity relationships (QSARs) of the enzymatic reactivity of the compounds, their electron-accepting potency and other reactivity indices were assessed by quantum mechanical methods. In P-450R-catalyzed reactions, both BFXs and NACs showed the same reactivity dependence on their electron-accepting potency which might be consistent with an “outer sphere” electron transfer mechanism. In NQO1-catalyzed two-electron (hydride) transferring reactions, BFXs acted as more efficient substrates than NACs, and the reduction efficacy of BFXs by NQO1 was in general higher than by single-electron transferring P-450R. In NQO1-catalyzed reactions, QSARs obtained showed that the reduction efficacy of BFXs, as well as that of NACs, was determined by their electron-accepting potency and could be influenced by their binding mode in the active center of NQO1 and by their global softness as their electronic characteristic. The reductive conversion of benzofuroxan by both flavoenzymes yielded the same reduction product of benzofuroxan, 2,3-diaminophenazine, with the formation of o-benzoquinone dioxime as a putative primary reductive intermediate, which undergoes a further reduction process. Overall, the data obtained show that by contrast to NACs, the flavoenzyme-catalyzed reduction of BFXs is unlikely to initiate their redox-cycling, which may argue for a minor role of the redox-cycling-type action in the cytotoxicity of BFXs. Full article