Abstract:

Phellodendron amurense is a traditional medicinal plant recognized for its abundant benzylisoquinoline alkaloids（BIAs）， showing substantial pharmacological properties. O-methyltransferases（OMTs）， pivotal enzymes in plant secondary metabolism， facilitate the methylation of phenolic hydroxyl groups and are crucial for the structural diversity and stability of BIAs. To comprehensively characterize the OMT gene family in P.amurense and identify key genes involved in BIA biosynthesis， this study performed a genome-wide analysis， identifying 82 OMT gene family members. Phylogenetic analysis indicated that all members belonged to the ClassⅠgroup， which can be further subdivided into two subgroups， ClassⅠa and ClassⅠb. Analysis of conserved motifs revealed that， while both subgroups maintained the core S-adenosylmethionine （SAM）-binding domain， they displayed notable differences in the composition of N- and C-terminal motifs. Chromosomal localization analysis illustrated the dispersed distribution of PaOMT genes. By integrating transcriptomic data from multiple tissues with in vitro enzymatic assays， PaOMT30 was identified to catalyze the O-methylation of the C-2 hydroxyl group of（S）-scoulerine， yielding（S）-tetrahydropalmatrubine. This study represented the first comprehensive analysis of the OMT gene family in P. amurense and successfully identified the C-2 methylation activity of PaOMT30 in BIA biosynthesis， establishing a foundation for elucidating the biosynthetic pathways of medicinal components in P.amurense and advancing BIA-based drug development.

Key words: Phellodendron amurense, benzylisoquinoline alkaloids, O-methyltransferase, methylation, functional characterization