NAD(P)H-(QUINONE-ACCEPTOR) OXIDOREDUCTASE OF TOBACCO-LEAVES IS A FLAVIN MONONUCLEOTIDE-CONTAINING FLAVOENZYME

The soluble NAD(P)H:(quinone-acceptor) oxidoreductase [NAD(P)H-QR, EC 1.6.99.2] of Nicotiana tabacum L. leaves and roots has been purified. NAD(P)H-QR contains noncovalently bound flavin mononucleotide. Pairs o f subunits of 21.4 kD are linked together by disulfide bridges, but th e active enzyme is a homotetramer of 94 to 100 kD showing an isoelectr ic point of 5.1. NAD(P)H-QR is a B-stereospecific dehydrogenase. NADH and NADPH are electron donors of similar efficiency with K-cat:K-m rat ios (with duroquinone) of 6.2 x 10(7) and 8.0 x 10(7) M(-1) s(-1), res pectively. Hydrophilic quinones are good electron accepters, although ferricyanide and dichlorophenolindophenol are also reduced. The quinon es are converted to hydroquinones by an obligatory two-electron transf er. No spectral evidence for a flavin semiquinone was detected followi ng anaerobic photoreduction. Cibacron blue and 7-iodo-acridone-4-carbo xylic acid are inhibitory. Tobacco NAD(P)H-QR resembles animal DT-diap horase in some respects (identical reaction mechanism with a two-elect ron transfer to quinones, unusually high catalytic capability, and don or and acceptor substrate specificity), but it differs from DT-diaphor ase in molecular structure, flavin cofactor, stereospecificity, and se nsitivity to inhibitors. As in the case with DT-diaphorase in animals, the main NAD(P)H-QR function in plant cells may be the reduction of q uinones to quinols, which prevents the production of semiquinones and oxygen radicals. The enzyme appears to belong to a widespread group of plant and fungal flavoproteins found in different cell compartments t hat are able to reduce quinones.