Identification of a novel multifunctional oxidosqualene cyclase from Zea mays sheds light on the biosynthetic pathway of three pentacyclic triterpenoids

Zea mays(maize)is an important agricultural crop that produces a variety of valuable terpenoids,including several triterpenoids.However,no oxidosqualene cyclase(OSC)enzymes,which catalyze the first step in triterpenoid biosynthesis,have been identified in maize.Here,we identified a novel OSC(ZmOSC1)in maize using a combination of genomic mining and phylogenetic analyses.To functionally characterize the candidate OSC,we constructed a yeast strain that produced high levels of 2,3-oxidosqualene.When ZmOSC1 was expressed in this strain,three compounds were detected and identified as hop-17(21)-en-3-ol,hopenol B and simiarenol,respectively.For their biosynthesis,we proposed a potential cyclization mechanism catalyzed by ZmOSC1 via the generation of a dammarenyl cation,followed by sequential cationic ring expansion,cyclization,cationic migration and further proton elimination.This study discovered and characterized an OSC from maize for the first time,and laid a foundation to produce three bioactive pentacyclic triterpenes,hop-17(21)-en-3-ol,hopenol B and simiarenol,using synthetic biology approaches.

Applications of protein engineering in the microbial synthesis of plant triterpenoids

Triterpenoids are a class of natural products widely used in fields related to medicine and health due to their biological activities such as hepatoprotection,anti-inflammation,anti-viral,and anti-tumor.With the advancement in biotechnology,microorganisms have been used as cell factories to produce diverse natural products.Despite the significant progress that has been made in the construction of microbial cell factories for the heterogeneous biosynthesis of triterpenoids,the industrial production of triterpenoids employing microorganisms has been stymied due to the shortage of efficient enzymes as well as the low expression and low catalytic activity of heterologous proteins in microbes.Protein engineering has been demonstrated as an effective way for improving the specificity,catalytic activity,and stability of the enzyme,which can be employed to overcome these challenges.This review summarizes the current progress in the studies of Oxidosqualene cyclases(OSCs),cytochrome P450s(P450s),and UDP-glycosyltransferases(UGTs),the key enzymes in the triterpenoids synthetic pathway.The main obstacles restricting the efficient catalysis of these key enzymes are analyzed,the applications of protein engineering for the three key enzymes in the microbial synthesis of triterpenoids are systematically reviewed,and the challenges and prospects of protein engineering are also discussed.