Diverse oxidosqualene cyclases for triterpenoid biosynthesis

Triterpenoids, a significant group of secondary metabolites, possess diverse skeletons and excellent pharmacological activities and are widely applied in cosmetics, medicine, and biopesticides. Traditional plant extraction and chemical synthesis cannot meet the needs of commercial production, and heterologous biosynthesis of triterpenoids has become a promising strategy. Oxidosqualene cyclases (OSCs) are the key enzymes that determine triterpenoid biosynthesis. However, most OSCs exhibit low heterologous expression, low catalytic activity, and promiscuous catalytic products, which impedes the heterologous biosynthesis of triterpenoids. In this study, we constructed a dataset of 348 OSCs with known functions, which is the largest centralized resource of characterized OSCs to date. The 88 triterpenoid skeletons produced by these OSCs are classified based on their structural features, 49 of which have been experimentally characterized as the main products of OSCs, and the formation mechanisms of the common skeletons are summarized. Phylogenetic analysis and motif analysis elucidated the relationships among the sequences, structures, and products of plant-derived OSCs. Phylogenetic analysis showed that plant-derived OSCs were divided into two major clades, which corresponded to the biosynthetic mechanisms of the "protosteryl/dammarenyl" dichotomy. Motif analysis showed that 5 motifs are present in plant-derived OSCs, including 3 related to the catalytic activity of the OSCs and 2 related to product specificity. Finally, we summarized the common strategies for mining OSCs to provide directions for the discovery of novel OSCs and new triterpenoid skeletons. Our study will help discover or design OSCs with specific product profiles as functional elements for applications in synthetic biology.