Purification and characterization of two forms of chicken liver cytochrome P-450 induced by 3,4,5,3',4'-pentachlorobiphenyl.
3,4,5,3',4'-Pentachlorobiphenyl (PenCB), one of the most potent 3-methylcholanthrene (MC)-type inducers of hepatic enzymes in animals, caused a remarkable induction of liver microsomal monooxygenases, particularly 7-ethoxyresorufin (7-ER) O-deethylase, benzo(a)pyrene (BP) 3-hydroxylase, and testosterone 16 alpha-hydroxylase in chickens, but not NADPH-cytochrome c(P-450) reductase and cytochrome b5. Two forms of cytochrome P-450 (P-450) in liver microsomes of PenCB-treated chickens were purified and characterized. The absorption maxima of the CO-reduced difference spectra of both enzymes (chicken P-448 L and chicken P-448 H) were at 448 nm. From the oxidized form of their absolute spectra, chicken P-448 L was a low-spin form and chicken P-448 H was a high-spin form. They had molecular masses of 56 and 54 kDa, respectively. In a reconstituted system, 7-ER O-deethylation, BP 3-hydroxylation, and testosterone 16 alpha-hydroxylation were catalyzed at high rates by chicken P-448 L but not by chicken P-448 H. Chicken P-448 L also catalyzed N-demethylation of aminopyrine, benzphetamine, and ethylmorphine with relatively low activity. On the other hand, chicken P-448 H functioned only in catalyzing estradiol 2-hydroxylation. These results were supported by an inhibition study of microsomal monooxygenases using an antibody against each enzyme. Immunochemical studies revealed that the enzymes differ from each other but are both inducible by PenCB-treatment. Chicken P-448 L and chicken P-448 H respectively comprise about 82 and 7% of the total P-450 content in chicken liver microsomes.
Hokama
Y
Y
Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka.
Koga
N
N
Yoshimura
H
H
D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D002645:Chickens; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004790:Enzyme Induction; D007120:Immunochemistry; D005779:Immunodiffusion; D007527:Isoenzymes / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009251:NADPH-Ferrihemoprotein Reductase / Q000302:isolation & purification; D011078:Polychlorinated Biphenyls / Q000494:pharmacology*; D011506:Proteins / Q000032:analysis; D039181:Steroid 16-alpha-Hydroxylase; D013379:Substrate Specificity
D000906:Antibodies; D007527:Isoenzymes; D011506:Proteins; D003577:Cytochrome P-450 Enzyme System; D011078:Polychlorinated Biphenyls; D039181:Steroid 16-alpha-Hydroxylase; D009251:NADPH-Ferrihemoprotein Reductase; C023035:3,4,5,3',4'-pentachlorobiphenyl