majority of cytochrome P450 enzymes (CYPs) predominantly operate as monooxygenases, but recently a class of P450 enzymes was discovered, that can act as peroxygenases (CYP152). The enzymes convert fatty acids through oxidative decarboxylation, yielding terminal alkenes, and through - and -hydroxylation to yield hydroxy-fatty acids. Bio-derived olefins may serve as biofuels and hence understanding the mechanism and substrate scope of this class of enzymes is important. In this work, we report on the substrate scope and catalytic promiscuity of CYP OleTJE and two of its orthologues from the CYP152 family, utilizing -monosubstituted branched carboxylic acids. We identify ,-desaturation as a dominant pathway for CYP OleTJE with 2-methylbutyric acid. To rationalize product distributions arising from /-hydroxylation, oxidative decarboxylation and desaturation depending on the substrate’s structure and binding pattern, a computational study was performed based on an active site complex of CYP OleTJE containing the heme cofactor in the substrate binding pocket and 2-methylbutyric acid as substrate. It is shown that substrate positioning determines the accessibility of the oxidizing species (Compound I) to the substrate and hence the regio- and chemo-selectivity of the reaction. Furthermore, the results show that for 2-methylbutyric acid, ,-desaturation is favorable because of a rate-determining -hydrogen atom abstraction, which cannot proceed to decarboxylation. Moreover, substrate hydroxylation is energetically impeded due to the tight shape and size of the substrate binding pocket.
- Cytochrome P450
- Density Functional Theory
- Valence Bond Modelling
- Reaction Mechanisms
Research Beacons, Institutes and Platforms
- Manchester Institute of Biotechnology