• BMB Reports
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• v.32 no.2
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• pp.127-132
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• 1999

The NAD(P)H-quinone oxidoreductase (EC 1. 6. 99. 2) was purified from S. cerevisiae. The native molecular weight of the enzyme is approximately 111 kDa and is composed of five identical subunits with molecular weights of 22 kDa each. The optimum pH of the enzyme is pH 6.0 with 1,4-benzoquinone as a substrate. The apparent $k_m$ for 1,4-benzoquinone and 1,4- naphthoquinone are 1.3 mM and $14.3\;{\mu}M$, respectively. Its activity is greatly inhibited by $Cu^{2+}$ and $Hg^{2+}$ ions, nitrofurantoin, dicumarol, and Cibacron blue 3GA. The purified NAD(P)H-quinone oxidoreductase was found capable of reducing aromatic nitroso compounds as well as a variety of quinones, and can utilize either NADH or NADPH as a source of reducing equivalents. The nitroso reductase activity of the purified NAD(P)H-quinone oxidoreductase is strongly inhibited by dicumarol.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.48-50
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• 2000

The energetics at the succinate:quinone oxidoreductase segment of V. alginolyticus was studied using a fluorescence quenching technique with inside-out membrane vesicles. A transient generation of the membrane potential (inside-positive) and ${\Delta}pH$ (inside-acidic) occurred in the presence of KCN and succinate when ubiquinone-1 (Q1) was added. The membrane potential (\Delta\psi$) generated by the succinate; quinone oxidoreductase segment was completely collapsed by the protonophore carbonylcyanide m-chlorophenylhydrazone (CCCP) and the membrane permeable anion $SCN^{-}$, whereas the ${\Delta}pH$ was completely collapsed by CCCP and $(NH_4)_2SO_4$. From these results, it was concluded that the succinate: quinone oxidoreductase segment as well as quinol oxidase [1] in the respiratory chain of V. alginolyticus generated $H^{+}$ electrochemical potential.

• BMB Reports
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• v.32 no.4
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• pp.321-325
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• 1999

NAD(P)H-quinone oxidoreductase (EC 1. 6. 99. 2) was purified form S. cerevisiae. The enzyme readily reduced 2,6-dichlorophenolindophenol, a quinonoid redox dye, as well as substituted benzo- and naphthoquinones, and could accept electrons from either NADH or NADPH. The purified NAD(P)H-quinone oxidoreductase turned out to be capable of reducing nitrosoarenes as well as a variety of quinones. A chemical-trapping technique using 4-chloro-1-naphthol was used to show that the N,N-dimethyl-p-benzoquinonediiminium cation was produced in the reduction of 4-nitroso-N,N-dimethylaniline catalyzed by NAD(P)H-quinone oxidoreductase.

• BMB Reports
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• v.40 no.1
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• pp.53-57
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• 2007

The enzymatic properties of NADH:quinone oxidoreductase were examined in Triton X-100 extracts of Bacillus cereus membranes by using the artificial electron acceptors ubiquinone-1 and menadione. Membranes were prepared from B. cereus KCTC 3674 grown aerobically on a complex medium and oxidized with NADH exclusively, whereas deamino-NADH was determined to be poorly oxidized. The NADH oxidase activity was lost completely by solubilization of the membranes with Triton X-100. However, by using the artificial electron acceptors ubiquinone-1 and menadione, NADH oxidation could be observed. The activities of NADH:ubiquinone-1 and NADH:menadione oxidoreductase were enhanced approximately 8-fold and 4-fold, respectively, from the Triton X-100 extracted membranes. The maximum activity of FAD-dependent NADH:ubiquinone-1 oxidoreductase was obtained at about pH 6.0 in the presence of 0.1M NaCl, while the maximum activity of FAD-dependent NADH:menadione oxidoreductase was obtained at about pH 8.0 in the presence of 0.1M NaCl. The activities of the NADH:ubiquinone-1 and NADH:menadione oxidoreductase were very resistant to such respiratory chain inhibitors as rotenone, capsaicin, and $AgNO_3$, whereas these activities were sensitive to 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO). Based on these results, we suggest that the aerobic respiratory chain-linked NADH oxidase system of B. cereus KCTC 3674 possesses an HQNO-sensitive NADH:quinone oxidoreductase that lacks an energy coupling site containing FAD as a cofactor.

• Journal of Life Science
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• v.24 no.1
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• pp.98-103
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• 2014

NAD(P)H quinone oxidoreductase 1 (NQO1) is a flavoprotein that catalyzes the two electron reduction of diverse substrates, including quinones. It uses NADH or NADPH as a cofactor for enzymatic machinery. In the metabolism of quinones, NQO1 has two conflicting functions because of the different stability of converted hydroquinones. The stable form of hydroquinone is excreted from cells by conjugation with glutathione or glucuronic acid. The unstable form of hydroquinone induces cell death by induction of oxidative stress and DNA damage. Certain quinones known as bio-reductive agents have a cytotoxic function following reduction by NQO1. Bio-reductive agents, such as ${\beta}$-lapachone or mitomycin C, induce the depletion of NAD(P)H and the generation of oxidative stress in an NQO1-dependent manner. NQO1 is highly expressed in several cancer tissues. Therefore, NQO1 is a good therapeutic target for cancer treatment with bio-reductive agents.

• BMB Reports
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• v.36 no.5
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• pp.442-449
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• 2003

NAD(P)H quinone oxidoreductase is a ubiquitous enzyme that is known to directly reduce quinone substrates to hydroquinones by a two-electron reaction. We report the identification of NADPH quinone oxidoreductase from Kluyveromyces marxianus (KmQOR), which reduces quinone substrates directly to hydroquinones. The KmQOR gene was sequenced, expressed in Escherichia coli, purified, and characterized. The open-reading frame of the KmQOR gene consists of 1143 nucleotides, encoding a 380 amino acid polypeptide. The nucleotide sequence of the KmQOR gene was assigned to EMBL under accession number AY040868. The $M_r$ that was determined by SDS-PAGE for the protein subunit was about 42 kDa, and the molecular mass of the native KmQOR was 84 kDa, as determined by column calibration, indicating that the native protein is a homodimer. The KmQOR protein efficiently reduced 1,4-benzoquinone, whereas no activities were found for menadiones and methoxyquinones. These observations, and the result of an extended sequence analysis of known NADPH quinone oxidoreductase, suggest that KmQOR possesses a different action mechanism.

• Archives of Pharmacal Research
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• v.24 no.5
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• pp.390-396
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• 2001

Synthesized 6-arylamino-5,8-quinolinediones 4a-4j and 6-chloro-7-arylamino-5,8-isoquinolinediones 5a-5g were evaluated for effects on NAD(P)H quinone oxidoreductase (NQOl ) activity with the cytosolic fractions derived from cultured human lung cancer cells and their cytotoxicity in cultured several human solid cancer cell lines. The 5,8-quinolinediones 4 and 5,8-isoquinolinediones 5 affected the reduction potential by NQO1 activity and showed a potent cytotoxic activity against human cancer cell lines. The tested compounds 4a, 5c, 5f, and 5g were considered as more potent cytotoxic agents. The compounds 4d, 5b, 5c, 5e and 5g were comparable modulators of NQO1 activity.

• Journal of Embryo Transfer
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• v.25 no.1
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• pp.35-42
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• 2010

Many studies propose that dysfunction of mitochondrial proton-translocating NADH-ubiquinone oxidoreductase (complex I) is associated with neurodegenerative disorders, such as Parkinson's disease and Huntington's disease. Mammalian mitochondrial proton-translocating NADH-quinone oxidoreductase (complex I) consists of at least 46 different subunits. In contrast, the NDI1 gene of Saccharomyces cerevisiae is a single subunit rotenone-insensitive NADH-quinone oxidoreductase that is located on the matrix side of the inner mitochondrial membrane. With a recombinant adeno-associated virus vector carrying the NDI1 gene (rAAV-NDI1) as the gene delivery method, we were able to attain high transduction efficiencies even in the human epithelial cervical cancer cells that are difficult to transfect by lipofection or calcium phosphate precipitation methods. Using a rAAV-NDI1, we demonstrated that the Ndi1 enzyme is successfully expressed in HeLa cells. The expressed Ndi1 enzyme was recognized to be localized in mitochondria by confocal immunofluorescence microscopic analyses and immunoblotting. Using digitonin-permeabilized cells, it was shown that the NADH oxidase activity of the NDI1-transduced HeLa cells were not affected by rotenone which is inhibitor of complex I, but was inhibited by flavone and antimycin A. The NDI1-transduced cells were able to grow in media containing rotenone. In contrast, control cells that did not receive the NDI1 gene failed to survive. In particular, in the NDI1-transduced cells, the yeast enzyme becomes integrated into the human respiratory chain. It is concluded that the NDI1 gene provides a potentially useful tool for gene therapy of mitochondrial diseases caused by complex I deficiency.

• Reproductive and Developmental Biology
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• v.35 no.4
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• pp.427-434
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• 2011

Mitochondria diseases have been reported to involve structural and functional defects of complex I-V. Especially, many of these diseases are known to be related to dysfunction of mitochondrial proton-translocating NADH-ubiquinone oxidoreductase (complex I). The dysfunction of mitochondria complex I is associated with neurodegenerative disorders, such as Parkinson's disease, Huntington's disease, and Leber's hereditary optic neuropathy (LHON). Mammalian mitochondrial proton-translocating NADH-quinone oxidoreductase (complex I) is largest and consists of at least 46 different subunits. In contrast, the NDI1 gene of Saccharomyces cerevisiae is a single subunit rotenone-insensitive NADH-quinone oxidoreductase that is located on the matrix side of the inner mitochondrial membrane. The Saccharomyces cerevisiae NDI1 gene using a recombinant adeno-associated virus vector (rAAV-NDI1) was successfully expressed in AML12 mouse liver hepatocytes and the NDI1-transduced cells were able to grow in media containing rotenone. In contrast, control cells that did not receive the NDI1 gene failed to survive. The expressed Ndi1 enzyme was recognized to be localized in mitochondria by confocal immunofluorescence microscopic analyses and immunoblotting. Using digitonin-permeabilized cells, it was shown that the NADH oxidase activity of the NDI1-transduced cells was not affected by rotenone which is inhibitor of complex I, but was inhibited by antimycin A. Furthermore, these results indicate that Ndi1 can be functionally expressed in the AML12 mouse liver hepatocytes. It is conceivable that the NDI1 gene is powerful tool for gene therapy of mitochondrial diseases caused by complex I deficiency. In the future, we will attempt to functionally express the NDI1 gene in mouse embryonic stem (mES) cell.

• Archives of Pharmacal Research
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• v.23 no.6
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• pp.554-558
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• 2000

Synthesized 5-arylamino-2-methyl-4,7-dioxobenzothiazoles 3a-3o were evaluated for modulation of NAD(P)H: quinone oxidoreductase (NQOl) activity with the cytosolic fractions derived from cultured human lung cancer cells and their cytotoxicity in cultured several human solid cancer cell lines. The 4,7-dioxobenzothiazoles affected the reduction potential by NQOl activity and showed a potent cytotoxic activity against human cancer cell lines. The tested compounds 3a, 3b, 3g, 3h, 3n and 3o were considered as more potent cytotoxic agents, and comparable modulators of NQOl activity.