Identification and characterization of three strictosidine synthases in Nauclea orientalis

Monoterpene indole alkaloids (MIAs) are a major class of plant-derived natural products with considerable clinical value. Nauclea orientalis accumulates diverse MIAs, yet their biosynthetic routes remain incompletely resolved. Here, by integrating transcriptome sequencing, phylogenetic analyses, and multiple-sequence alignments, we identified five candidates underpinning the formation of the universal MIA precursor strictosidine: one loganic acid O-methyltransferase (NoLAMT1), one secologanin synthase (NoSLS1), and three strictosidine synthases (NoSTR1-3). Comprehensive in vitro and in planta assays demonstrated that NoLAMT1 methylates loganic acid to yield loganin, which is subsequently oxidized by NoSLS1 to secologanin; NoSTR1-3 then catalyze the Pictet-Spengler condensation of secologanin with tryptamine to produce strictosidine. These findings shed light on the molecular basis of strictosidine formation in N. orientalis and furnish tractable genetic parts for heterologous reconstruction and sustainable MIA production.