Characterization of FMN-dependent NADH-quinone reductase induced by menadione in Escherichia coli.

It was found that when Escherichia coli is grown in the presence of 0.2-0.3 mM menadione (2-methyl-1,4-naphthoquinone), an FMN-dependent NADH-quinone reductase increases more than 20-fold in the cytoplasmic fraction. The menadione-induced quinone reductase was isolated from the cytoplasmic fraction of induced cells. The purified enzyme had an Mr of 24 kDa on SDS-polyacrylamide gel electrophoresis. The enzyme required flavin as a cofactor and a half-maximum activity was obtained with 0.54 microM FMN or 16.5 microM FAD. The enzyme had a broad pH optimum at pH 7.0-8.0 and reacted with NADH, but not with NADPH. The reaction followed a ping-pong mechanism and the intrinsic Km values for NADH and menadione were estimated to be 132 microM and 2.0 microM, respectively. Dicoumarol was a simple competitive inhibitor with respect to NADH with a Ki value of 0.22 microM. The electron acceptor specificity of this enzyme was very similar to that of NAD(P)H: (quinone acceptor) oxidoreductase (EC 1.6.99.2, DT-diaphorase) from rat liver. Since menadione is reduced by the two-electron reduction pathway to menadiol, the induction of this enzyme is likely to be an adaptive response of E. coli to partially alleviate the toxicity of menadione.