Structure-function relationship of NAD(P)H:quinone reductase: characterization of amino-terminal blocking group and essential tyrosine and lysine residues

Haniu, Mitsuru; Yuan, Henry; Chen, Shiuan; Iyanagi, Takashi; Lee, Terry D.; Shively, John E.

doi:10.1021/bi00418a033

Cited by 26 publications

(11 citation statements)

References 27 publications

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“…Its amino acid sequence has been determined (Haniu et al, 1988) and is in agreement with that deduced from the cDNA sequence (Robertson et al, 1986;Bayney et al, 1987). Extensive structurefunction studies of the rat enzyme have been performed by affinity and photoaffinity labeling techniques (Haniu et al, 1988;Liu et al, 1989;Deng et al, 1991) and site-directed mutagenesis experiments (Forrest et al, 1990;Chen et al, 1992;Ma et al, 1992). Site-directed mutagenesis experiments were carried out using either COS cell-or Escherichia coli-expression systems.…”

supporting

confidence: 71%

“…This peptide has a sequence homologous with that of the amino-terminus of rat quinone reductase that was also found to be blocked by an acetyl group (Haniu et al, 1988), leading to the hypothesis that this peptide is the amino-terminal peptide of mouse quinone reductase. This was confirmed when the complete amino acid sequence was generated by cloning (see below).…”

Section: Characterization Of the Amino-terminal Peptide By Mwms Analysismentioning

confidence: 99%

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Mouse liver NAD(P)H:quinone acceptor oxidoreductase: Protein sequence analysis by tandem mass spectrometry, cDNA cloning, expression in Escherichia coli, and enzyme activity analysis

et al. 1994

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The amino acid sequence of mouse liver NAD(P)H:quinone acceptor oxidoreductase (EC 1.6.99.2) has been determined by tandem mass spectrometry and deduced from the nucleotide sequence of the cDNA encoding for the enzyme. The electrospray mass spectral analyses revealed, as previously reported (Prochaska HJ, Talalay P, 1986, J Biol Chem 261: [1372][1373][1374][1375][1376][1377][1378], that the 2 forms -the hydrophilic and hydrophobic forms-of the mouse liver quinone reductase have the same molecular weight. No amino acid sequence differences were found by tandem mass spectral analyses of tryptic peptides of the 2 forms. Moreover, the amino-termini of the mouse enzymes are acetylated as determined by tandem mass spectrometry. Further, only 1 cDNA species encoding for the quinone reductase was found. These results suggest that the 2 forms of the mouse quinone reductase have the same primary sequences, and that any difference between the 2 forms may be attributed to a labile posttranslational modification.Analysis of the mouse quinone reductase cDNA revealed that the enzyme is 273 amino acids long and has a sequence homologous to those of rat and human quinone reductases. In this study, the mouse quinone reductase cDNA was also ligated into a prokaryotic expression plasmid pKK233.2, and the constructed plasmid was used to transform Escherichia coli strain JM109. The E. coli-expressed mouse quinone reductase was purified and characterized. Although mouse quinone reductase has an amino acid sequence similar to those of the rat and human enzymes, the mouse enzyme has a higher NAD(P)H-menadione reductase activity and is less sensitive to flavones and dicoumarol, 2 known inhibitors of the enzyme. The results would indicate that the regions in mouse quinone reductase that contain amino acids different from the rat and human enzymes are critical for the binding of menadione, flavones, and dicoumarol.

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supporting

confidence: 71%

Section: Characterization Of the Amino-terminal Peptide By Mwms Analysismentioning

confidence: 99%

Mouse liver NAD(P)H:quinone acceptor oxidoreductase: Protein sequence analysis by tandem mass spectrometry, cDNA cloning, expression in Escherichia coli, and enzyme activity analysis

et al. 1994

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“…4. It builds on the features of the previously proposed NAD(P)H binding sites (8,10) and suggests that two quinone molecules are reduced to hydroquinones by accepting electrons from NAD(P)H at both subunits (Fig. 4).…”

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confidence: 57%

Subunit functional studies of NAD(P)H:quinone oxidoreductase with a heterodimer approach.

Cui

Yang

1995

Proc. Natl. Acad. Sci. U.S.A.

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Studies on the active sites of the homodimer proteins by construction of hybrids of wild-type and site-directed mutant proteins have been reported (13-15). Wente and Schachman (13) used an approach which entails the formation of such hybrids in vitro, but this approach is only applicable to proteins which can sustain the dissociation and reassociation conditions. Larimer et al. (14) reported the production of a hybrid heterodimer ofwild type and a site-directed mutant by plasmid cotransformation. Nevertheless, the study was limited by the fact that the three species of dimeric proteins (wild-type homodimer, mutant homodimer, heterodimer) could not be separated from one another.In the present study, we have developed an approach to express a heterodimer (H194A/HNQOR) containing a wildtype subunit with a polyhistidine tag at the amino terminus of NQOR and a H194A mutant subunit by constructing two cDNAs under one promoter in Escherichia coli. This approach enabled us to separate heterodimer from the two homodimers by stepwise elution with imidazole from a nickel nitrilotriacetate (Ni-NTA) column under nondenaturing conditions. The functions of the heterodimer subunits were studied by characterizing the kinetics of the reduction of 2,6-dichloroindophenol (DCIP), menadione, and methyl red.

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“…30, No. 28, 1991 6943 Wistar rats injected daily for 3 days with 3-methylcholanthrene (4 mg/100 g of body weight), according to a procedure described by Haniu et al (1988).…”

Section: Methodsmentioning

confidence: 99%

“…The primary structure of the rat liver NAD(P)H:quinone acceptor reductase has been deduced from cDNA sequences (Robertson et al, 1986;Bayney et al, 1987) and determined by protein microsequencing methods (Haniu et al, 1988). The amino acid sequence of the human enzyme has also been deduced from the cDNA sequence (Jaswal et al, 1988).…”

mentioning

confidence: 99%

Photodependent inhibition of rat liver NAD(P)H:quinone acceptor oxidoreductase by (A)-2-azido-NAD+ and (A)-8-azido-NAD+

Deng¹,

Zhao²,

Iyanagi³

et al. 1991

Biochemistry

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Two photoaffinity analogues of NAD+, (A)-2-azido-NAD+ [nicotinamide 2-azidoadenine dinucleotide] and (A)-8-azido-NAD+ [nicotinamide 8-azidoadenine dinucleotide], have been synthesized, and their reactivities with the rat liver NAD(P)H:quinone acceptor oxidoreductase have been investigated. The reduce nicotinamide nucleotide probes, (A)-2-azido-NADH and (A)-8-azido-NADH, were shown to be substrates of the quinone reductase. This enzyme was inhibited by (A)-8-azido-NADH, were shown to be substrates of the quinone reductase. This enzyme was inhibited by (A)-2-azido-NAD+ and (A)-8-azido-NAD+ in a photodependent manner, and the inhibition of the enzyme could be prevented by the presence of nicotinamide nucleotide substrates during photolysis. (A)-2-Azido-NAD+ was demonstrated to be a more potent inhibitor than (A)-8-azido-NAD+. In addition, the photodependent inhibition by (A)-8-azido-NAD+ increased when menadione, the substrate of the enzyme, was present during the photolysis, while menadione protected the enzyme from the photodependent inhibition by (A)-2-azido-NAD+. These results indicate that these two NAD+ analogues can be used to identify the nicotinamide nucleotide binding site of this quinone reductase and that they probably bind to the enzyme in different fashions.

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Structure-function relationship of NAD(P)H:quinone reductase: characterization of amino-terminal blocking group and essential tyrosine and lysine residues

Cited by 26 publications

References 27 publications

Mouse liver NAD(P)H:quinone acceptor oxidoreductase: Protein sequence analysis by tandem mass spectrometry, cDNA cloning, expression in Escherichia coli, and enzyme activity analysis

Mouse liver NAD(P)H:quinone acceptor oxidoreductase: Protein sequence analysis by tandem mass spectrometry, cDNA cloning, expression in Escherichia coli, and enzyme activity analysis

Subunit functional studies of NAD(P)H:quinone oxidoreductase with a heterodimer approach.

Photodependent inhibition of rat liver NAD(P)H:quinone acceptor oxidoreductase by (A)-2-azido-NAD+ and (A)-8-azido-NAD+

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