INTERACTION OF POLYCYCLIC AROMATIC-HYDROCARBONS AND FLAVONES WITH CYTOCHROMES P450 IN THE ENDOPLASMIC-RETICULUM - EFFECT ON CO BINDING-KINETICS

The flash photolysis technique was used to examine the kinetics of CO binding to cytochromes P450 in rat liver microsomes. The effect of pol ycyclic aromatic hydrocarbons (PAHs) and flavones was used to distingu ish the kinetic behavior of the PAH-metabolizing P450 1A1 from that of the remaining multiple microsomal P450s. Applying this approach to mi crosomes from 3-methylcholanthrene-treated rats showed that although a ll tested PAHs accelerated CO binding to P450 1A1, the extent varied m arkedly for different PAHs. The tricyclic PAHs phenanthrene and anthra cene enhanced CO binding by 37- and 49-fold, respectively, while sever al tetracyclic and pentacyclic PAHs increased the rate by 3-16-fold. T he results indicate that PAHs exert a dual effect on the rate of CO bi nding to P450 1A1: a general enhancement via widening of the CO access channel and a reduction that is dependent on PAH size. Although 5,6-b enzoflavone increased the rate of CO binding to P450 1A1 by 3.5-fold, it additionally decelerated binding to a constitutive P450 by 15-fold. This flavone thus exerts markedly different effects on two P450s with in the same microsomal sample. In contrast, the sole effect of 7,8-ben zoflavone was acceleration of CO binding to P450 1A1 by 18-fold. The d ivergent effects of these isomeric flavones, which only differ in posi tioning of an aromatic ring, illustrate the sensitivity of CO binding to substrate structure. The varying effects of these PAHs and flavones on CO binding kinetics show that they differentially modulate P450 co nformation and access of ligands to the P450 heme and demonstrate that binding of carcinogens to a specific target P450 can be evaluated in its native microsomal milieu.