Induction by Triacetyloleandomycin and partial purification of a LM 3 form of cytochrome P-450 from rabbit liver microsomes

Abstract

Treatment of rabbits with Triacetyloleandomycin (a currently used antibiotic in human therapy) at 1 mmol per kg of body weight daily for 5 days results in a significant induction of liver microsomal cytochrome P-450, (2.6 nmol/mg proteins). Electrophoresis in SDS polyacrylamide gels shows this increase in P-450 is associated to the appearance of a strong band in a zone located between the major bands of microsomes induced by phenobarbital and β-naphtoflavone (LM 3 forms in Coon's terminology). Partial purification of this P-450 LM 3 (TAO) was undertaken by chromatographic procedures (CM cellulose and hydroxylapatite). Its subunit molecular weight is 52 000; the absolute spectra in the oxidized, ferrous and CO-ferrous forms present maxima at 417, 536, and 570 nm; 415 and 548 nm; 450 and 555 nm respectively. Monooxygenase activity of LM 3 (TAO) was compared with that of LM 2 and LM 4 in a reconstituted system containing NADPH cytochrome P-450 reductase and phosphatidylcholine; the activity of P-450 LM 3 (TAO) was higher than that of LM 2 and LM 4 with chlorcyclizine as a substrate. According to these observations, LM 3 (TAO) resembles LM 3 (b), a constitutive form of untreated rabbit liver microsomes.