A UDP-rhamnosyltransferase (PviUGT74) in Panax vietnamensis var. fuscidiscus completing pseudoginsenoside F11 biosynthesis

Pseudoginsenoside F 11 (PF 11 ), a rare ocotillol-type saponin with neuroprotective and anti-ischemic properties. The crucial rhamnosyltransferase, for its final glycosylation step has yet to be identified. This study conducted a comprehensive genome-wide analysis of the UGT family in Panax vietnamensis var. fuscidiscus and performed in vitro enzyme activity validation, identifying the UDP-rhamnosyltransferase PviUGT74, which can catalyze the C6- O -rhamnosylation of PRT 5 to produce PF 11 . The enzyme demonstrated optimal activity at near-neutral pH and moderate temperature, with strict specificity for UDP-rhamnosyltransferase. Homologs from six other Panax species exhibited similar activity, indicating strong evolutionary conservation. Molecular docking and site-directed mutagenesis identified seven critical residues (H21, D120, E273, W355, G356, Y375, E376) essential for catalysis. Synteny analysis suggested that PviUGT74 arose from transpositional duplication, followed by neofunctionalization. This study clarifies the final enzymatic step of PF 11 biosynthesis and lays a mechanistic and biotechnological foundation for its sustainable production through engineered biosynthesis, minimizing dependence on plant extraction. • A total of 110 PviUGT genes were identified in the genome of Panax vietnamensis var. fuscidiscus . • PviUGT74 uses UDP-rhamnose as a sugar donor to catalyze C-6 glycosylation of PRT 5 , forming PF 11 . • Homologous of PviUGT74 were identified in six Panax species, and functional validation confirmed retained catalytic activity. • Molecular docking and alanine scanning experiments identified key amino acid residues for the enzyme's catalytic activity.