CATALYTIC ACTIVITY OF CYTOCHROME P4501A2 IN RECONSTITUTED SYSTEM WITHEMULGEN 913

Physicochemical properties and catalytic activity of cytochrome P4501A 2 in the reconstituted system with Emulgen 913 were studied. The forma tion of cytochrome P4501A2 monomers was shown by gel filtration at an Emulgen concentration of 8 g/liter. The catalytic activity of the mono meric monooxygenase system was low. The maximum rate of the 7-ethoxyre sorufin O-deethylation reaction, 150 pmol resorufin min(-1) nmol(-1) c ytochrome P4501A2, was observed at an Emulgen concentration of 0.1 g/l iter when cytochrome P4501A2 pentamers predominated. The effects of Em ulgen on cytochrome P4501A2 cumene hydroperoxide-dependent peroxidase activity, on its affinity to substrate and to cytochrome b(5), and on the rate constants of dithionite-dependent reduction were insignifican t. Study of the NADPH-dependent reduction of cytochrome P4501A2 in the reconstituted system showed that the rate constant and reduction leve l of cytochrome P4501A2 were always higher when the reaction was initi ated by NADPH than when it was initiated by NADPH-cytochrome P450 redu ctase. This indicated that the reduction reaction initiated by the red uctase was limited by a step in the cytochrome P4501A2 and reductase i nteraction. Correlation between 7-ethoxyresorufin O-deethylase activit y and reduction level of cytochrome P4501A2 was found. (C) 1994 Academ ic Press, Inc.