A prostaglandin ω-hydroxylase cytochrome P-450 (P-450(PG-ω)) purified from lungs of pregnant rabbits

D. E. Williams, S. E. Hale, R. T. Okita, B. S.S. Masters

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82 Scopus citations

Abstract

Cytochrome P-450-dependent prostaglandin ω-hydroxylation is induced over 100-fold during late gestation in rabbit pulmonary microsomes Purification of cytochromes P-450 from lung microsomes of pregnant rabbits yielded three fractions. Two of these fractions correspond to rabbit lung P-450(I) (LM2) and P-450(II) (LM5), which together constitute 70-97% of total cytochrome P-450 in lung microsomes from nonpregnant rabbits. The third form, which we designate rabbit cytochrome P-450(PG-ω), regioselectively hydroxylates prostaglandins at the ω-position in reconstituted systems with a turnover of 1-5 min-1. Titration with purified pig liver cytochrome b5, demonstrated a 4-fold maximum stimulation at a cytochrome b5 to a P-450 molar ratio. of 1-2. Rabbit lung P-450(PG-ω) formed a typical type I binding spectrum upon the addition of prostaglandin E1 with a calculated K(s) of 1 μM, which agreed reasonably well with the kinetically calculated K(m) of 3 μM. Cytochrome P-450(PG-ω) was isolated as a low-spin isozyme with a λ(max) (450 nm) in the CO-difference spectrum distinguishable from P-450(I) (451 nm) and P-450(II) (449 nm). Sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis demonstrated that although purified P-450(PG-ω) had a relatively low specific content (12.1 nmolmg-1), it appeared homogeneous with a calculated minimum M(r) of 56,000, intermediate between rabbit LM4 and LM6. When lung microsomes from pregnant and nonpregnant rabbit were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a protein band, with a M(r) identical to P-450(PG-ω), was observed in the pregnant rabbit, whereas this band appeared to be very faint or absent in microsomes from the nonpregnant rabbit. Purification of cytochromes P-450 from nonpregnant rabbit lung yielded only P-450(I) and P-450(II). P-450(PG-ω) appears to be a novel rabbit P-450, possessing high activity towards ω-hydroxylation of prostaglandins, and is greatly induced during pregnancy in rabbit lung.

Original languageEnglish (US)
Pages (from-to)14600-14608
Number of pages9
JournalJournal of Biological Chemistry
Volume259
Issue number23
StatePublished - 1984

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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