Functional Expression of House Fly (Musca domestica) Cytochrome P450 CYP6D1 in Yeast (Saccharomyces cerevisiae)

Cytochrome P450 CYP6D1 from the house fly is important in the detoxication of xenobiotics and in resistance to pyrethroid insecticides. In house fly microsomes CYP6D1 requires cytochrome b₅ for the metabolism of some substrates, such as benzoa]pyrene, but does not require cytochrome b₅ for the metabolism of other substrates such as methoxyresorufin. To examine the molecular mechanisms involved in its metabolism of pyrethroids and other substrates, a system for the heterologous expression of CYP6D1 in the yeast Saccharomyces cerevisiae was developed. Heterologous CYP6D1 can be inducibly expressed by culture in media with galactose as the sole carbon source, and is successfully inserted into the yeast microsomes. CYP6D1 is enzymatically active, as measured by methoxyresorufin-O-demethylation, indicating that CYP6D1 is able to interact with yeast P450 reductase. However, CYP6D1 expression did not result in measurable benzoa]pyrene hydroxylation, suggesting that CYP6D1 cannot interact with yeast cytochrome b₅, or that there is insufficient cytochrome b₅ in the yeast microsomes to support this CYP6D1-mediated activity. Some suggestions are made for improving the yeast microsomal oxidoreductase environment in order to optimize CYP6D1 function.