Cytochromes P450 (CYPs) catalyse diverse reactions and are key enzymes in fungal primary and secondary metabolism, and xenobiotic detoxification. CYP enzymatic properties and substrate specificity determine the reaction outcome. However, CYP-mediated reactions may also be influenced by their redox partners. Filamentous fungi with numerous CYPs often possess multiple microsomal redox partners, cytochrome P450 reductases (CPRs). In the plant pathogenic ascomycete Cochliobolus lunatus we recently identified two CPR paralogues, CPR1 and CPR2. Our objective was to functionally characterize two endogenous fungal cytochrome P450 systems and elucidate the putative physiological roles of CPR1 and CPR2. We reconstituted both CPRs with CYP53A15, or benzoate 4-hydroxylase from C. lunatus, which is crucial in the detoxification of phenolic plant defence compounds. Biochemical characterization using RP-HPLC shows that both redox partners support CYP activity, but with different product specificities. When reconstituted with CPR1, CYP53A15 converts benzoic acid to 4-hydroxybenzoic acid, and 3-methoxybenzoic acid to 3-hydroxybenzoic acid. However, when the redox partner is CPR2, both substrates are converted to 3,4-dihydroxybenzoic acid. Deletion mutants and gene expression in mycelia grown on media with inhibitors indicate that CPR1 is important in primary metabolism, whereas CPR2 plays a role in xenobiotic detoxification.
COBISS.SI-ID: 4729882
Grosmannia clavigera is a fungal associate of the mountain pine beetle (Dendroctonus ponderosae) and a pathogen of lodgepole pine (Pinus contorta) that must overcome terpenoid oleoresin and phenolic defenses of host trees. G. clavigera responds to monoterpene influx with complementary mechanisms that include export and the use of these compounds as a carbon source. Cytochromes P450 (CYPs) may also be involved in the metabolism of host defense compounds. We have identified and phylogenetically classified G. clavigera CYPs (CYPome). We show that although the G. clavigera CYPome has contracted in evolution, certain CYP families have expanded by duplication. We analyzed RNA-seq data for CYP expression following treatment with terpenes and pine phloem extracts to identify CYPs potentially involved in detoxification of these pine defense compounds. We also used transcriptome analysis of G. clavigera grown on monoterpenes, triglycerides or oleic acid as a carbon source to identify up-regulated CYPs that may be involved in the utilization of these compounds to support fungal growth. Finally, we identify secondary metabolite biosynthetic gene clusters that contain CYPs, and CYPs in clusters that may be involved in conversion of host chemicals.
COBISS.SI-ID: 30257113