The use of Yarrowia lipolytica for the expression of human cytochrome P450 CYP1A1.
Cytochromes P450 constitute a superfamily of haem-thiolate mono-oxygenases that are involved in the oxidative metabolism of lipophilic subtrates. These enzymes require association with cytochrome P450 reductase ( CPR) to achieve optimal activities. We have expressed human cytochrome P450 CYP1A1 under the POX2 promoter (pPOX2-CYP1A1) in Y. lipolytica, with or without overproduction of Y. lipolytica CPR expressed under the ICL promoter (pICL- CPR) or the POX2 promoter (pPOX2- CPR). Activity of cytochrome CYP1A1 was analysed by conversion of hydroxyresorufin to resorufin. Strain JMY330 and JMY330- CPR present no activity, the monocopy cytochrome CYP1A1 integrant JMY331 and JMY331- CPR derivatives present an average activity of 32.0 pM/min/dw and 48.3 and 64.6 pM/min/dw for pICL- CPR and pPOX2- CPR, respectively. Increase of CPR expression resulted in about two-fold higher activity. The multicopy 1A1 integrant JMY339 and JMY339- CPR derivatives present an activity of 129 pM/min/dw and 815-1845 pM/min/dw, respectively. Increase of CPR expression resulted in 6.3-12.8-fold higher activity, depending on the CPR transformant. We observed a 50-fold increase of activity between the monocopy integrant JMY331 as compared to the multicopies integrant JMY339- CPR in which CPR was overexpressed.[1]References
- The use of Yarrowia lipolytica for the expression of human cytochrome P450 CYP1A1. Nthangeni, M.B., Urban, P., Pompon, D., Smit, M.S., Nicaud, J.M. Yeast (2004) [Pubmed]
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