Affinity modification of microsomal flavoproteins by NAD(P) 2',3'-dialdehydes

NADPH-cytochrome P-450 reductase (FP1) and NADH-cytochrome b5 reductase (FP2) involved in the microsomal fraction of rat liver have been modified chemically by periodate-oxidized NADP+ and NAD+ (o-NAD(P]. Despite its low Ki values (approximately 30 microM) o-NADP is not covalently bound with FP1, although o-NAD with Ki greater than 100 microM chemically modifies FP1 by suppressing its activity. The protective effect of NADP+ against FP1 inactivation indicates that FP1 is modified in the NADP+ binding site. An active centre of FP2 is modified by o-NAD in the same manner as FP1 (NAD+ prevents FP2 from inactivation). FP2 is slightly inactivated when the concentration of o-NADP is one order of magnitude higher than that of o-NAD. As found, the o-NAD-modified microsomal FP1 inhibits the oxidation of cytochrome P-450 substrates (acetanilide and p-nitroanisole).