Increased TCA cycle activity and reduced oxygen consumption during cytochrome P450-dependent biotransformation in fission yeast

Cytochrome P450s are haern-containing monooxygenases that catalyse a variety of oxidations utilizing a large substrate spectrum and are therefore of interest for biotechnological applications. We expressed human CYP21 in fission yeast Schizosaccharomyces pombe as a eukaryotic model for P450-dependent whole-cell biotransformation. The resulting strain displayed strong steroid hydroxylase activity that was accompanied by contrary effects on respiration and non-respiratory oxygen consumption, which combined to a significant decline in total oxygen consumption of the cells. While production of ROS (reactive oxygen species) decreased, the TCA cycle activity increased, as was shown by metabolic flux (METAFoR) analysis. Pentose phosphate pathway (PPP) activity was found to be negligible, regardless of growth phase, CYP21 expression or biocatallytic activity, indicating that NADPH levels in Sz. poinbe are sufficiently high to support an exogenous P450 without adaptations of central carbon metabolism. We conclude from these data that neither oxygen supply nor NADPH availability are limiting factors in'P450-dependent biocatalysis in Sz. pombe. Copyright (c) 2006 John 'Wiley & Sons, Ltd.