The diterpenoid paclitaxel(Taxol)is a chemotherapy medication widely used as a first-line treatment against several types of solid cancers.The supply of paclitaxel from natural sources is limited.However,missing knowledge about the genes involved in several specific metabolic steps of paclitaxel biosynthesis has rendered it difficult to engineer the full pathway.In this study,we used a combination of transcriptomics,cell biology,metabolomics,and pathway reconstitution to identify the complete gene set required for the het-erologous production of paclitaxel.We identified the missing steps from the current model of paclitaxel biosynthesis and confirmed the activity of most of the missing enzymes via heterologous expression in Nico-tiana benthamiana.Notably,we identified a new C4β-C20 epoxidase that could overcome the first bottleneck of metabolic engineering.We used both previously characterized and newly identified oxomutases/epoxi-dases,taxane 1 β-hydroxylase,taxane 9α-hydroxylase,taxane 9α-dioxygenase,and phenylalanine-CoA ligase,to successfully biosynthesize the key intermediate baccatin Ⅲ and to convert baccatin Ⅲ into pacli-taxel in N.benthamiana.In combination,these approaches establish a metabolic route to taxoid biosynthesis and provide insights into the unique chemistry that plants use to generate complex bioactive metabolites.
展开▼
