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Purification and Quantification of Saccharomyces cerevisiae Cytochrome P450

  • Ian Stansfield
  • Steven L. Kelly
Part of the Methods in Molecular Biology™ book series (MIMB, volume 53)

Abstract

Cytochrome P450s (cyt.P450s) are a class of hemoprotein enzymes found in a wide variety of organisms, where they play an important role in endogenous metabolism and the metabolism of xenobiotic compounds. In eukaryotes these enzymes are membrane-bound, in most cases located in the endoplasmic reticulum. Higher eukaryotes often have complex systems containing many cyt.P450s. However, vegetatively growing Saccharomyces cerevisiae contains only one major cyt.P450, the lanosterol 14α demethylase (1), and thus provides an ideal model system for the study of microsomal eukaryote cyt.P450s. Methods will be described for the quantification of yeast cyt.P450s, and the purification of the main yeast cyt.P450. This latter method was first described by Yoshida and Aoyama (1), and King et al. (2). Recently, the mRNA of a second S. cerevisiae cyt.P450 gene has been detected among sporulation-specific transcripts (3).

Keywords

Ammonium Sulfate Microsomal Protein Sodium Dithionite Sodium Cholate Microsome Preparation 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Yoshida, Y. and Aoyama, Y. (1984) Yeast cytochrome P450 catalysing lanosterol 14α demethylation. I. Purification and spectral properties. J. Biol. Chem. 259, 1655–1660.PubMedGoogle Scholar
  2. 2.
    King, D. J., Azari, M. R., and Wiseman, A. (1984) Studies on the properties of highly purified cytochrome P448 and its dependent benzo[a]pyrene hydroxylase from Saccharomyces cerevisiae. Xenobiotica 14, 187–206.PubMedCrossRefGoogle Scholar
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    Briza, P., Breitenbach, M., Ellinger, A., and Segall, J. (1991) Isolation of two developmentally regulated genes involved in spore maturation in Saccharomyces cerevisiae. Genes Dev. 4, 1775–1789.CrossRefGoogle Scholar
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    Ching, M. S., Lennard, M. S., Tucker, G. T., Woods, H. F., Kelly, D. E., and Kelly, S. L. (1991) Expression of human cytochrome P450 1A1 in the yeast Saccharomyces cerevisiae. Biochem. Pharmacol. 42, 753–758.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1996

Authors and Affiliations

  • Ian Stansfield
    • 1
  • Steven L. Kelly
    • 2
  1. 1.Biological LaboratoryUniversity of KentCanterburyUK
  2. 2.Krebs Institute for Biomolecular ResearchUniversity of SheffieldUK

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