Unexpected genetic and structural relationships of a long-forgotten flavoenzyme to NAD(P)H:quinone reductase (DT-diaphorase)

Zhao, Qinjian; Yang, Xiao; Holtzclaw, W. David; Talalay, Paul

doi:10.1073/pnas.94.5.1669

Cited by 119 publications

(111 citation statements)

References 29 publications

Supporting

Mentioning

105

Contrasting

Order By: Relevance

“…All other chemicals were of the highest purity commercially available. Reduction of N-(2-hydroxylethylnicotinamide) to N-(2-hydroxylethyldihydronicotinamide) was carried out just before use by sodium hydrosulfite (19).…”

Section: Chemicals Reagents and Antibodiesmentioning

confidence: 99%

See 1 more Smart Citation

Crystal Structure of Quinone Reductase 2 in Complex with Resveratrol^,

et al. 2004

View full text Add to dashboard Cite

Resveratrol has been shown to have chemopreventive, cardioprotective, and antiaging properties. Here, we report that resveratrol is a potent inhibitor of quinone reductase 2 (QR2) activity in vitro with a dissociation constant of 35 nM and show that it specifically binds to the deep active-site cleft of QR2 using high-resolution structural analysis. All three resveratrol hydroxyl groups form hydrogen bonds with amino acids from QR2, anchoring a flat resveratrol molecule in parallel with the isoalloxazine ring of FAD. The unique active-site pocket in QR2 could potentially bind other natural polyphenols such as flavonoids, as proven by the high affinity exhibited by quercetin toward QR2. K562 cells with QR2 expression suppressed by RNAi showed similar properties as resveratrol-treated cells in their resistance to quinone toxicity. Furthermore, the QR2 knockdown K562 cells exhibit increased antioxidant and detoxification enzyme expression and reduced proliferation rates. These observations could imply that the chemopreventive and cardioprotective properties of resveratrol are possibly the results of QR2 activity inhibition, which in turn, upregulates the expression of cellular antioxidant enzymes and cellular resistance to oxidative stress.Resveratrol (trans-3,4′,5-trihydroxystilbene) is a phyto-polyphenol that occurs in grapes and a variety of medicinal plants. Because of its abundance in grapes, it is present in significant amount in grape products such as grape juice and wines, particularly red wine (1). The revealing of resveratrol as a cancer chemopreventive agent in 1997 (2) sparked extensive research on its chemopreventive properties (3). Resveratrol has been tested on a variety of cancers in animal model studies. As in the case of the 1997 study, Jang et al. (2) demonstrated in a mouse model that resveratrol is effective in prevention of DMBA-(dimethylbenzanthracene) and TPA-(tetradecanoylphorbol-13-acetate) induced skin cancer. Application of 1, 5, 10, or 25 µM of resveratrol with TPA twice a week for 18 weeks reduced the number of skin tumors per mouse by 68, 81, 76, or 98%, respectively, and the percentage of mice with tumors was lowered by 50, 63. 63, or 88%. In mouse models of colon cancer and small intestinal cancer, resveratrol prevents cancer formation with 100 and 70% efficacy, respectively (4, 5). † This work was supported in part by National Institute of Health Grant R21 CA104424. ‡ The atomic coordinates and structure factors (PDB code 1SG0) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptResveratrol has been suggested to block the multistep process of carcinogenesis, namely, tumor initiation, promotion, and progression (3). Despite extensive investigation, no clear molecular basis for the actions of resveratrol has emerged. A variety of hypotheses have been proposed to explain its functions: antioxidant effects, proapoptotic effects, cell-cycle regulation, inhibition of protein kinases, regulation of NFκB and Iκ3, and modulation of estrogen effects (3, 6). ...

show abstract

Section: Chemicals Reagents and Antibodiesmentioning

confidence: 99%

“…The snugness of the fit of resveratrol into the cleft suggests that longer molecules or those with a nonflat conformation will not fit into the cavity. This explains why QR1 inhibitors such as dicumarol and cibacron blue do not potently inhibit QR2 activity (19).…”

Section: Structure Of the Qr2-resveratrol Complexmentioning

confidence: 99%

Crystal Structure of Quinone Reductase 2 in Complex with Resveratrol^,

et al. 2004

View full text Add to dashboard Cite

show abstract

“…9; access number JO2888). Liver mRNA (200 ng) was reverse-transcribed with oligo(dT) [12][13][14][15][16][17][18] in accordance with the firststrand cDNA synthesis protocol from Amersham Pharmacia Biotech. Polymerase chain reactions were performed in 100 l containing 10 mM Tris-HCl, pH 8.3, 1.5 mM MgCl 2 , 0.2 mM dNTP, 2 l of the singlestranded cDNA preparation, 0.3 M of each primer, and 2 units of pfu native polymerase (Stratagene) with a 35 cycles program of 94°C for 1 min, 65°C for 2 min, and 72°C for 2 min and a final extension at 72°C for 8 min.…”

Section: Cdna-derived Expression Of Hqr 2 In Hamster Ovary Cho Cells mentioning

confidence: 99%

“…It is noteworthy that the literature on QR 2 enzymology is rather scarce (12)(13)(14). Interestingly, QR 2 was originally discovered in 1962 as a flavoenzyme (12), later re-discovered as the QR 1 -related enzyme (14), and was recently found again in porcine kidney as a puromycin aminonucleosidebinding protein (15). We now unveiled a new facet of QR 2 as the melatonin-binding site MT 3, opening new perspectives in melatonin investigations as well as in quinone reduction studies.…”

mentioning

confidence: 99%

Identification of the Melatonin-binding SiteMT 3 as the Quinone Reductase 2

Nosjean¹,

Ferro²,

Cogé³

et al. 2000

Journal of Biological Chemistry

540

358

View full text Add to dashboard Cite

The regulation of the circadian rhythm is relayed from the central nervous system to the periphery by melatonin, a hormone synthesized at night in the pineal gland. Besides two melatonin G-coupled receptors, mt 1 and MT 2 , the existence of a novel putative melatonin receptor, MT 3 , was hypothesized from the observation of a binding site in both central and peripheral hamster tissues with an original binding profile and a very rapid kinetics of ligand exchange compared with mt 1 and MT 2 . In this report, we present the purification of MT 3 from Syrian hamster kidney and its identification as the hamster homologue of the human quinone reductase 2 (QR 2 , EC 1.6.99.2). Our purification strategy included the use of an affinity chromatography step which was crucial in purifying MT 3 to homogeneity. The protein was sequenced by tandem mass spectrometry and shown to align with 95% identity with human QR 2 . After transfection of CHO-K1 cells with the human QR 2 gene, not only did the QR 2 enzymatic activity appear, but also the melatonin-binding sites with MT 3 characteristics, both being below the limit of detection in the native cells. We further confronted inhibition data from MT 3 binding and QR 2 enzymatic activity obtained from samples of Syrian hamster kidney or QR 2 -overexpressing Chinese hamster ovary cells, and observed an overall good correlation of the data. In summary, our results provide the identification of the melatonin-binding site MT 3 as the quinone reductase QR 2 and open perspectives as to the function of this enzyme, known so far mainly for its detoxifying properties.Melatonin, a neurohormone produced at night in the pineal gland, is suspected to relay to the peripheral organs the circadian rhythm detected by the central nervous system. Several high affinity melatonin receptors have been identified to date, among which the mt 1 (1) and MT 2 (2) receptors have been cloned from human tissues. The pharmacology of these two receptors is well documented, and several compounds, including melatonin, are ligands with picomolar binding affinity (for review, see Ref.3). Another putative melatonin receptor was identified on pharmacological grounds, with lower melatonin affinity (nanomolar range), very rapid ligand association/dissociation kinetics, and an original pharmacological profile (4 -6). In line with mt 1 and MT 2 receptors, this putative receptor was named MT 3 , according to the nomenclature recommendations of the IUPHAR (7). So far, the known inhibitors of MT 3 hardly reach the nanomolar range and encompass an unusually large structural diversity of highly hydrophobic cyclic or polycyclic compounds (Refs. 5 and 6, and for review, see Ref.3).1 All pharmacological investigations on mt 1 , MT 2 , and MT 3 were performed using the radioligand [125 I]melatonin, a ligand with high affinity for mt 1 and MT 2 (K d ϭ 10 -200 pM) and with lower affinity for MT 3 (K d ϭ 3-9 nM). The hamster kidney, liver, and brain have been used as model tissues for MT 3 pharmacological studies, and our recent data con...

show abstract

“…3). NADH-dh appeared to be different from those flavoproteins classified in EC 1.1.99.2 [4][5][6] and EC 1.6.99. 3,7,8) most of which are unable to use oxygen as the electron y To whom correspondence should be addressed.…”

mentioning

confidence: 99%