Molecular Pharmacology, Vol 15, 708-718, Copyright © 1979 by the American Society for Pharmacology and Experimental Therapeutics

Multiple Forms of Cytochrome P-450: Catalytic Differences Exhibited by Two Homogeneous Forms of Rabbit Cytochrome P-450

¹ Department of Biochemistry, Scripps Clinic and Research Foundation, La Jolla, California 92037

Two highly purified forms of hepatic cytochrome P-450 were isolated from rabbits treated with the inducers phenobarbital and 2,3,7,8-tetrachlorodibenzo-p-dioxin. Both forms were shown to be metabolically active when reconstituted with lipid and homogeneous NADPH-cytochrome P-450 reductase. Form 2, obtained from rabbits treated with phenobarbital, and form 4, purified from rabbits treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin, hydroxylated biphenyl at comparable rates (V_max) 4.2 and 3.4 mol min^-1 mol^-1 cytochrome P-450, respectively, although with dissimilar apparent K_m values, 31 µM and 9.6 µM, respectively. Form 4 was 60 times more active than form 2 in the deethylation of 7-ethoxyresorufin. Moreover, form 4 was five orders of magnitude more sensitive in the biphenyl reaction to the inhibitory effects of the differential inhibitor -naphthoflavone than was form 2. Densitometric scans of sodium dodecyl sulfate polyacrylamide gel electrophoretograms revealed that microsomes from rabbits treated with phenobarbital exhibit a relative predominance of form 2. In similar fashion, microsomes from 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated animals show a dominance of form 4. The metabolic activities of the induced microsomes reflect the influence of the major form of cytochrome P-450. Similar trends are seen in the activities of phenobarbital-induced microsomes and reconstituted form 2 just as 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced microsomes and reconstituted form 4 share like metabolic properties. Thus, the distinct catalytic capacities of multiple forms of cytochrome P-450 represent significant determinants of xenobiotic metabolism by the cytochrome P-450 system.

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ACKNOWLEDGMENTS We thank Ms. Maryann C. Zounes for her technical assistance in providing the SDS PAGE, and Dr. Richard L. Norman for providing the densitometric scans for the SDS PAGE.

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