Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.5.2 (NQO1)
 6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mammalian NAD(P)H:quinone oxidoreductase (NQO1, DT-diaphorase, EC 1.6.99.2) catalyzes the two-electron reduction of quinones and plays one of the main roles in the bioactivation of quinoidal drugs. In order to understand the enzyme substrate specificity, we have examined the reactions of rat NQO1 with a number of quinones with available potentials of single-electron (E(1)(7)) reduction and pK(a) of their semiquinones. The hydride transfer potentials (E(7)(H(-))) were calculated from the midpoint potentials of quinones and pK(a) of hydroquinones. Our findings imply that benzo- and naphthoquinones with a van der Waals volume (VdWvol) < or = 200 A(3) are much more reactive than glutathionyl-substituted naphthoquinones, polycyclic quinones, and FMN (VdWvol>200 A(3)) with the same reduction potentials. The entropies of activation (DeltaS(not equal)) in the reduction of "fast" oxidants are equal to -84 to -76 J mol(-1) K(-1), whereas in the reduction of "slow" oxidants Delta S(not equal)=-36 to -11 J mol(-1) K(-1). The large negative Delta S(not equal) in the reduction of fast oxidants may be explained by their better electronic coupling with reduced FAD or the formation of charge-transfer complexes, since fast oxidants bind at the dicumarol binding site, whereas the binding of some slow oxidants outside it has been demonstrated. The reactivity of quinones may be equally well described in terms of the three-step (e(-),H(+),e(-)) hydride transfer, using E(1)(7), pK(a)(QH*), and VdWvol as correlation parameters, or in terms of single-step (H(-)) hydride transfer, using E(7)(H(-)) and VdWvol in the correlation. The analysis of NQO1 reactions with single-electron acceptors and quinones using an "outer-sphere" electron transfer model points to the possibility of a three-step hydride transfer.
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PMID:Two-electron reduction of quinones by rat liver NAD(P)H:quinone oxidoreductase: quantitative structure-activity relationships. 1214 63

NQO1 (DT-diaphorase) and its truncated isoenzyme, the metalloenzyme NQO2, can reduce quinone substrates by two-electron transfer. While NQO1 is a known detoxification enzyme, the function of NQO2 is less well understood. Both rat NQO1 and human NQO2 reductively bioactivate the dinitroarene CB 1954 to a cytotoxic product that behaves as a difunctional DNA-crosslinking species with potent anti-tumour activity, although human NQO1 is much less effective. A FMN-dependent nitroreductase from E. coli B also reduces quinones and reductively bioactivates CB 1954. However, this enzyme reduces CB 1954 to the 2- and 4-hydroxylamines in equivalent yield, whereas NQO1 and NQO2 generate only the 4-isomer. The reduction profile is a key factor in the development of anti-tumour prodrugs, where distinct delivery strategies are being evaluated: prodrug therapy, antibody-, macromolecule and gene-directed enzyme prodrug therapy (ADEPT, MDEPT or GDEPT). The flavoprotein enzymes are explored in terms of structure and bioreduction mechanism, particularly for use in the design of novel prodrugs with potential application as chemotherapeutic agents.
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PMID:Aerobic nitroreduction by flavoproteins: enzyme structure, mechanisms and role in cancer chemotherapy. 1236 76

Acryloyl-CoA reductase from Clostridium propionicum catalyses the irreversible NADH-dependent formation of propionyl-CoA from acryloyl-CoA. Purification yielded a heterohexadecameric yellow-greenish enzyme complex [(alpha2betagamma)4; molecular mass 600 +/- 50 kDa] composed of a propionyl-CoA dehydrogenase (alpha2, 2 x 40 kDa) and an electron-transferring flavoprotein (ETF; beta, 38 kDa; gamma, 29 kDa). A flavin content (90% FAD and 10% FMN) of 2.4 mol per alpha2betagamma subcomplex (149 kDa) was determined. A substrate alternative to acryloyl-CoA (Km = 2 +/- 1 microm; kcat = 4.5 s-1 at 100 microm NADH) is 3-buten-2-one (methyl vinyl ketone; Km = 1800 microm; kcat = 29 s-1 at 300 microm NADH). The enzyme complex exhibits acyl-CoA dehydrogenase activity with propionyl-CoA (Km = 50 microm; kcat = 2.0 s-1) or butyryl-CoA (Km = 100 microm; kcat = 3.5 s-1) as electron donor and 200 microm ferricenium hexafluorophosphate as acceptor. The enzyme also catalysed the oxidation of NADH by iodonitrosotetrazolium chloride (diaphorase activity) or by air, which led to the formation of H2O2 (NADH oxidase activity). The N-terminus of the dimeric propionyl-CoA dehydrogenase subunit is similar to those of butyryl-CoA dehydrogenases from several clostridia and related anaerobes (up to 55% sequence identity). The N-termini of the beta and gamma subunits share 40% and 35% sequence identities with those of the A and B subunits of the ETF from Megasphaera elsdenii, respectively, and up to 60% with those of putative ETFs from other anaerobes. Acryloyl-CoA reductase from C. propionicum has been characterized as a soluble enzyme, with kinetic properties perfectly adapted to the requirements of the organism. The enzyme appears not to be involved in anaerobic respiration with NADH or reduced ferredoxin as electron donors. There is no relationship to the trans-2-enoyl-CoA reductases from various organisms or the recently described acryloyl-CoA reductase activity of propionyl-CoA synthase from Chloroflexus aurantiacus.
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PMID:Acryloyl-CoA reductase from Clostridium propionicum. An enzyme complex of propionyl-CoA dehydrogenase and electron-transferring flavoprotein. 1260 23

Inclusion of an oligomeric enzyme, NAD+-dependent hydrogenase from the hydrogen-oxidizing bacterium Ralstonia eutropha, into a system of reverse micelles of different sizes resulted in its dissociation into catalytically active heterodimers and subunits, which were characterized in reactions with various substrates. It was found that: 1) the native tetrameric form of this enzyme catalyzes all types of studied reactions; 2) hydrogenase dimer, HoxHY, is a minimal structural unit catalyzing hydrogenase reaction with an artificial electron donor, reduced methyl viologen; 3) all structural fragments containing FMN and NAD+/NADH-binding sites exhibit catalytic activity in diaphorase reactions with one- and two-electron acceptors; 4) small subunits, HoxY and HoxU also exhibit activity in diaphorase reactions with artificial acceptors. These results can be considered as indirect evidence that the second FMN molecule may be associated with one of the small subunits (HoxY or HoxU) of the hydrogenase from R. eutropha.
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PMID:Use of a reverse micelle system for study of oligomeric structure of NAD+-reducing hydrogenase from Ralstonia eutropha H16. 1603 6

The crystal structure of AzoR (azoreductase) has been determined in complex with FMN for two different crystal forms at 1.8 and 2.2 A resolution. AzoR is an oxidoreductase isolated from Escherichia coli as a protein responsible for the degradation of azo compounds. This enzyme is an FMN-dependent NADH-azoreductase and catalyzes the reductive cleavage of azo groups by a ping-pong mechanism. The structure suggests that AzoR acts in a homodimeric state forming the two identical catalytic sites to which both monomers contribute. The structure revealed that each monomer of AzoR has a flavodoxin-like structure, without the explicit overall amino acid sequence homology. Superposition of the structures from the two different crystal forms revealed the conformational change and suggested a mechanism for accommodating substrates of different size. Furthermore, comparison of the active site structure with that of NQO1 complexed with substrates provides clues to the possible substrate-binding mechanism of AzoR.
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PMID:Three-dimensional structure of AzoR from Escherichia coli. An oxidereductase conserved in microorganisms. 1668 76

In order to identify an enzyme capable of Fenton reaction in Synechocystis, we purified an enzyme catalyzing one-electron reduction of t-butyl hydroperoxide in the presence of FAD and Fe(III)-EDTA. The enzyme was a 26 kDa protein, and its N-terminal amino acid sequencing revealed it to be DrgA protein previously reported as quinone reductase [Matsuo M, Endo T and Asada K (1998) Plant Cell Physiol39, 751-755]. The DrgA protein exhibited potent quinone reductase activity and, furthermore, we newly found that it contained FMN and highly catalyzed nitroreductase, flavin reductase and ferric reductase activities. This is the first demonstration of nitroreductase activity of DrgA protein previously identified by a drgA mutant phenotype. DrgA protein strongly catalyzed the Fenton reaction in the presence of synthetic chelate compounds, but did so poorly in the presence of natural chelate compounds. Its ferric reductase activity was observed with both natural and synthetic chelate compounds with a better efficiency with the latter. In addition to small molecular-weight chemical chelators, an iron transporter protein, transferrin, and an iron storage protein, ferritin, turned out to be substrates of the DrgA protein, suggesting it might play a role in iron metabolism under physiological conditions and possibly catalyze the Fenton reaction under hyper-reductive conditions in this microorganism.
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PMID:Synechocystis DrgA protein functioning as nitroreductase and ferric reductase is capable of catalyzing the Fenton reaction. 1729 43

The crystal structure of the flavodoxin-like protein WrbA with oxidized FMN bound reveals a close relationship to mammalian NAD(P)H:quinone oxidoreductase, Nqo1. Structural comparison of WrbA, flavodoxin, and Nqo1 indicates how the twisted open-sheet fold of flavodoxins is elaborated to form multimers that extend catalytic function from one-electron transfer between protein partners using FMN to two-electron reduction of xenobiotics using FAD. The structure suggests a novel physiological role for WrbA and Nqo1.
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PMID:WrbA bridges bacterial flavodoxins and eukaryotic NAD(P)H:quinone oxidoreductases. 1789 67

Two high-molecular-weight rubredoxin genes (hrb) from Clostridium thermocellum and Moorella thermoacetica were expressed in Escherichia coli and their translated products (Hrb) were characterized. M. thermoacetica Hrb showed strong diaphorase activity. In contrast, C. thermocellum Hrb, containing neither FMN nor FAD in the molecule, showed flavin reductase activity with moderate diaphorase activity. Both enzymes were optimally active at about 50 degrees C.
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PMID:Two proteins with diaphorase activity from Clostridium thermocellum and Moorella thermoacetica. 1832 61

A small enzyme showing diaphorase activity was purified from culture supernatant of Clostridium kluyveri and its N-terminal amino acid sequence was determined. This sequence identified a gene (diaA) encoding a protein (DiaA) of 229 amino acids with a predicted molecular weight of 24,981 in the genomic DNA sequence database of C. kluyveri constructed by the Research Institute of Innovative Technology for the Earth. The predicted protein was composed of a flavin reductase-like domain and a rubredoxin-like domain from its N-terminus. The diaA gene was cloned into an expression vector and expressed in an Escherichia coli recombinant. Recombinant enzyme rDiaA showed NADH/NADPH diaphorase activity with 2,6-dichlorophenolindophenol and nitro blue tetrazolium. The enzyme was most active at pH 8.0 at 40 degrees C. The UV-visible absorption spectrum and thin layer chromatography (TLC) analyses indicated that one rDiaA molecule contained a tightly bound FMN molecule as a prosthetic group. An iron molecule was also detected in an enzyme molecule.
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PMID:Cloning and expression of a Clostridium kluyveri gene responsible for diaphorase activity. 1832 62

Analysis of the amino acid sequences of subunits NuoM and NuoN in the membrane domain of Complex I revealed a clear common pattern, including two lysines that are predicted to be located within the membrane, and which are important for quinone reductase activity. Site-directed mutations of the amino acid residues E144, K234, K265 and W243 in this pattern were introduced into the chromosomal gene nuoM of Escherichia coli Complex I. The activity of mutated Complex I was studied in both membranes and in purified Complex I. The quinone reductase activity was practically lost in K234A, K234R and E144A, decreased in W243A and K265A but unchanged in E144D. Complex I from all these mutants contained 1 mol tightly bound ubiquinone per mol FMN like wild type enzyme. The mutant enzymes E144D, W243A and K265A had wild type sensitivity to rolliniastatin and complete proton-pumping efficiency of Complex I. Remarkably, the subunits NuoL and NuoH in the membrane domain also appear to contain conserved lysine residues in transmembrane helices, which may give a clue of the mechanism of proton translocation. A tentative principle of proton translocation by Complex I is suggested based on electrostatic interactions of lysines in the membrane subunits.
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PMID:Conserved lysine residues of the membrane subunit NuoM are involved in energy conversion by the proton-pumping NADH:ubiquinone oxidoreductase (Complex I). 1859 Jun 97


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