CATALYTIC PROPERTIES OF NAD(P)H-QUINONE ACCEPTOR OXIDOREDUCTASE - STUDY INVOLVING MOUSE, RAT, HUMAN, AND MOUSE-RAT CHIMERIC ENZYMES

NAD(P)H:quinone acceptor oxidoreductase (quinone reductase) (DT-diapho rase, EC 1.6.99.2) is involved in the process of reductive activation of cytotoxic antitumor quinones and nitrobenzenes. In this study, we i nitially examined the relative abilities of mouse, rat, and human quin one reductases to reduce two prodrugs, CB 1954 [5-(aziridin-1-yl)-2,4- dinitrobenzamide] and EO9 nyl)-3-(hydroxymethyl)-2-(3-hydroxy-1-propen yl)-1- methyl-1H-indole-4,7-dione]. By using Escherichia coil-expresse d quinone reductases and evaluating them under identical conditions, w e confirmed previous findings showing that the human enzyme is not as effective as the rat enzyme in reducing CB 1954 and EO9, although the two enzymes have similar NAD(P)H-menadione reductase activities. Inter estingly, although the amino acid sequence of mouse quinone reductase is more homologous to that of the rat enzyme, we found that the mouse enzyme behaves similarly to the human enzyme in its ability to reduce these compounds and to generate drug-induced DNA damage. To determine the region of quinone reductase that is responsible for the catalytic differences, two mouse-rat chimeric enzymes were generated, MR-P, a ch imeric enzyme that has mouse amino-terminal and rat carboxyl-terminal segments of quinone reductase, was shown to have catalytic properties resembling those of rat quinone reductase, and RM-P, a chimeric enzyme that has rat amino-terminal and mouse carboxyl-terminal segments of q uinone reductase, was shown to have catalytic properties resembling th ose of mouse quinone reductase. In addition, MR-P and RM-P were found to be inhibited by flavones with K-l values similar to those for rat a nd mouse quinone reductases, respectively. Based on these results, we propose that the carboxyl-terminal portion of the enzyme plays an impo rtant role in the reduction of cytotoxic drugs and the binding of flav ones.