A modular UDP-glycosyltransferase and cytochrome P450 system for soyasaponin C-22 modification.

Soybean (Glycine max (L.) Merr.) produces triterpenoid saponins, called soyasaponins, which are categorised as 2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP) saponins and group A saponins based on the chemical modifications at the C-22 position of their triterpene skeleton. These C-22 modifications confer different biological functions. While most genes involved in soyasaponin biosynthesis have been identified, the enzymes responsible for conjugation of DDMP or arabinose to the C-22 hydroxyl group remain unclear. This study aimed to identify the enzymes involved in these modifications. Co-expression analysis using known saponin biosynthetic genes identified three candidate UGT genes. Among them, UGT73K5 was essential for DDMP saponin biosynthesis in soybean, as ugt73k5 mutants lacked these compounds. Similarly, the orthologous gene in Lotus japonicus, UGT73K6, was required for DDMP saponin biosynthesis, and the ugt73k6 defect was complemented by soybean UGT73K5. In addition, UGT73BU1, a homologous gene not co-expressed with known saponin biosynthetic genes, was identified as a C-22 arabinosyltransferase involved in group A saponin biosynthesis. In L. japonicus, UGT73K6 is genomically co-localised and co-expressed with a cytochrome P450 gene responsible for C-22 hydroxylation, forming a conserved biosynthetic gene cluster. This organisation suggests coordinated biosynthetic modules underlying DDMP saponin production in legumes.