Immunochemical characterization of a cytochrome P450 isozyme and a protein purified from liver microsomes of male guinea pigs and their roles in the oxidative metabolism of Δ9-tetrahydrocannabinol by guinea pig liver microsomes

Shizuo Narimatsu, Yuko Akutsu, Tamihide Matsunaga, Kazuhito Watanabe, Ikuo Yamamoto, Hidetoshi Yoshimura

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A protein (designated as protein-B) was purified from liver microsomes of adult male guinea pigs by an affinity chromatography with ω-aminooctyl Sepharose 4B, followed by HPLC using DEAE-5PW and hydroxyapatite columns which had been used to purify a cytochrome P450 (P450) isozyme (P450-A) from the same subcellular fraction (Narimatsu et al., Biochem Biophys Res Commun 172: 607-613, 1990). Protein-B had a molecular mass of 49 kDa in SDS-PAGE, but did not show absorbance at 417 nm for heme. Further, it did not show any oxidative activities towards aniline (AN), d-benzphetamine (d-BP), p-nitroanisole (p-NA) or Δ9-tetrahydrocannabinol (Δ9-THC) in a reconstituted system including dilauroylphosphatidylcholine, NADPH-P450 reductase, and cytochrome b5. However, antiserum against protein-B raised in rabbits suppressed liver microsomal oxidative activities towards d-BP and p-NA dose-dependently. The antibody decreased Δ9-THC oxidation activity most effectively, but did not decrease AN hydroxylation activity. Antiserum against P450-A suppressed all the activities towards these four substrates, especially towards Δ9-THC, in liver microsomes of male guinea pigs. Moreover, reconstitution with hemin made it possible for protein-B to produce some oxidative activity toward Δ9-THC. These results suggest that protein-B is also a cytochrome P450 isozyme which has lost a heme moiety during purification steps. Both P450-A and protein-B could have a role as cytochrome P450 isozymes in the oxidative metabolism of drugs, especially that of Δ9-THC by the liver microsomes of adult male guinea pigs.

Original languageEnglish
Pages (from-to)1729-1736
Number of pages8
JournalBiochemical Pharmacology
Issue number8
Publication statusPublished - Apr 15 1992


ASJC Scopus subject areas

  • Biochemistry
  • Pharmacology

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