Purification and characterization of an intracellular NADH: quinone reductase from Trametes versicolor.
Intracellular NADH:quinone reductase involved in degradation of aromatic compounds including lignin was purified and characterized from white rot fungus Trametes versicolor. The activity of quinone reductase was maximal after 3 days of incubation in fungal culture, and the enzyme was purified to homogeneity using ion-exchange, hydrophobic interaction, and gel filtration chromatographies. The purified enzyme has a molecular mass of 41 kDa as determined by SDS-PAGE, and exhibits a broad temperature optimum between 20-40 degrees C , with a pH optimum of 6.0. The enzyme preferred FAD as a cofactor and NADH rather than NADPH as an electron donor. Among quinone compounds tested as substrate, menadione showed the highest enzyme activity followed by 1,4-benzoquinone. The enzyme activity was inhibited by CuSO(4), HgCl(2), MgSO(4), MnSO(4), AgNO(3), dicumarol, KCN, NaN(3), and EDTA. Its Km and Vmax with NADH as an electron donor were 23 microM and 101 mM/mg per min, respectively, and showed a high substrate affinity. Purified quinone reductase could reduce 1,4-benzoquinone to hydroquinone, and induction of this enzyme was higher by 1,4-benzoquinone than those of other quinone compounds.
Lee
Sang-Soo
SS
Division of Life Sciences, and Research Institute of Life Sciences, Kangwon National University, Chuncheon 200-701, Republic of Korea.
Moon
Dong-Soo
DS
Choi
Hyoung T
HT
Song
Hong-Gyu
HG
D016227:Benzoquinones / Q000378:metabolism; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006873:Hydroquinones / Q000378:metabolism; D008670:Metals / Q000494:pharmacology; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*; D020072:Polyporales / Q000201:enzymology*; D013053:Spectrophotometry; D013696:Temperature
D016227:Benzoquinones; D005656:Fungal Proteins; D006873:Hydroquinones; D008670:Metals; C004532:quinone; D009249:NADP; D004492:Edetic Acid; D016660:NAD(P)H Dehydrogenase (Quinone); C031927:hydroquinone