Format

Send to

Choose Destination
Biochimie. 1990 Jun-Jul;72(6-7):463-72.

Maximizing the expression of mammalian cytochrome P-450 monooxygenase activities in yeast cells.

Author information

1
Centre de Génétique Moléculaire du CNRS, Laboratoire Propre associé à l'Université Pierre-et-Marie-Curie, Gif-sur-Yvette, France.

Abstract

Cytochrome P-450s constitute a superfamily of mono-oxygenases which require the association with specific redox enzymes bound to the endoplasmic reticulum membrane for their activity. Conditions for the functional expression of these mammalian enzymes in yeast cells and the respective merits and limitations of currently used P-450 expression systems, are considered. The dependence of the mouse P-450 IA1 specific activity on the cytochrome expression level in yeast microsomes is studied and results demonstrate that the low amounts of endogenous NADPH-cytochrome P-450 reductase and cytochrome b5 which are naturally present, are limiting for the heterologous monooxygenase activities. The sequences encoding human liver cytochrome b5, the native and a modified form of the yeast NADPH-cytochrome P-450 reductase were cloned by making use of PCR techniques, over-expressed in yeast as functional forms, and characterized. New vectors allowing a high level of mammalian P-450 expression upon induction were also constructed and tested. A strategy for the construction of a co-expression system allowing maximal activity of mammalian cytochrome P-450s is discussed.

PMID:
2124149
DOI:
10.1016/0300-9084(90)90070-w
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Elsevier Science
Loading ...
Support Center