The ancient CYP716 family is a major contributor to the diversification of eudicot triterpenoid biosynthesis

Abstract

Triterpenoids are widespread bioactive plant defence compounds with potential use as pharmaceuticals, pesticides and other high-value products. Enzymes belonging to the cytochrome P450 family have an essential role in creating the immense structural diversity of triterpenoids across the plant kingdom. However, for many triterpenoid oxidation reactions, the corresponding enzyme remains unknown. Here we characterize CYP716 enzymes from different medicinal plant species by heterologous expression in engineered yeasts and report ten hitherto unreported triterpenoid oxidation activities, including a cyclization reaction, leading to a triterpenoid lactone. Kingdom-wide phylogenetic analysis of over 400 CYP716s from over 200 plant species reveals details of their evolution and suggests that in eudicots the CYP716s evolved specifically towards triterpenoid biosynthesis. Our findings underscore the great potential of CYP716s as a source for generating triterpenoid structural diversity and expand the toolbox available for synthetic biology programmes for sustainable production of bioactive plant triterpenoids.

Multi‐tissue integration of transcriptomic and specialized metabolite profiling provides tools for assessing the common bean (Phaseolus vulgaris) metabolome

“…Saponin‐related CYPs included previously characterized CYP93E9 (Phvul.008G128700.1) (Moses et al ., ) involved in β‐amyrin hydroxylation and several other genes divided in four main clades comprising proteins from the CYP72A and CYP716A families, as well as two smaller clades formed with GuCYP88D6 and MtCYP87D16. The CYP716A family was recently shown to be a major contributor to triterpenoid diversity, being conserved in eudicots and involved in the biosynthesis of a variety of different aglycones (Miettinen et al ., ). Even though there are no saponin‐related CYP716A members characterized in soybean or common bean, we could identify three genes (Phvul.002G302100, Phvul.008G034400 and Phvul.008G034500) exhibiting high similarity to CYP716A12 and CYP716A179 from Medicago and G. uralensis , respectively.…”

Comparative analysis of CYP716A subfamily enzymes for the heterologous production of C-28 oxidized triterpenoids in transgenic yeast

“…Subsequently, cytochrome P450 enzymes modify these triterpenols, yielding the corresponding C-28 oxidized products. Recently, several C-28 oxidizing enzymes have been identified in various plant species (Carelli et al 2011;Fukushima et al 2011;Miettinen et al 2017;Seki et al 2015). The industrial production of C-28 oxidized products is an active research area (Li and Zhang 2015;Zhou et al 2016).…”

Multi‐omics data‐driven investigations of metabolic diversity of plant triterpenoids

“…Members from the subfamilies CYP51G, CYP85A, CYP90B‐D, CYP710A, CYP724B and CYP734A that play essential roles in modification of the tetracyclic scaffolds to generate phytosterols or phytohormones are generally conserved in all plant species, while P450s from CYP51, CYP72 and CYP85 clans are recruited from the primary metabolic pathway and have evolved new functions for triterpene metabolism (Hamberger and Bak, ). For instance, the CYP716 family from the CYP85 clan is conserved across land plants but has evolved specifically toward pentacyclic triterpene metabolism in eudicots (Miettinen et al ., ). CYP716As are conserved in higher plants and catalyze a consecutive three‐step oxidation at C28 of α‐amyrin, lupel and β‐amyrin (Carelli et al ., ; Fukushima et al ., ; Misra et al ., ; Tamura et al ., ).…”