Expression, purification and charactization of cytochrome P450 4A4

A. E. Aitken, L. J. Roman, B. S.S. Masters

Research output: Contribution to journalArticlepeer-review

Abstract

Members of the cytochrome P450 4A subfamily catalyze the ohydroxylation of prostaglandins, fatty acids and eicosanoids. Cytochrome P450 4A4 (4A4), a member of this subfamily, was first purified from pregnant rabbit lung (Williams et al, 1984) and concurrently from progesterone-treated male rabbits (Yamamoto e( a/, 1984). The induction of 4A4 in pregnant rabbit lung suggests that it is of physiological importance although the precise function is not known. Previously, this laboratory has expressed cytochrome P450 4A4 in E. coli (Nishimoto et al, 1993) but low yields have limited the usefulness of this system. An improved E. coli expression and purification system has been developed. This system provides increased yields of pure detergent-free cytochrome P450 4A4(30-50 nmole liter1). Current work has involved the kinetic characterization of this expressed protein by examination of activity with its preferred substrate prostaglandin E, (Km = 10M; Turnover No. = 9.9min'). This is a 2-fold increase in activity compared to the previously expressed cytochrome P450 4A4 (Nishimoto ef a/, 1993).

Original languageEnglish (US)
Pages (from-to)A804
JournalFASEB Journal
Volume11
Issue number9
StatePublished - Dec 1 1997

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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