Catalytic divergence of O‐methyltransferases shapes the chemo‐diversity of polymethoxylated bibenzyls in Dendrobium catenatum

• Published in: The Plant journal : for cell and molecular biology Vol. 120; no. 1; pp. 29 - 44
• Main Authors: Ta, He, Yang, Ya‐Hui, Zhu, Ting‐Ting, Du, Ni‐Hong, Hao, Yue, Fu, Jie, Xu, Dan‐Dan, Xu, Ze‐Jun, Cheng, Ai‐Xia, Lou, Hong‐Xiang
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01.10.2024
• Subjects: Bioaccumulation; biochemical pathways; Biosynthesis; biosynthetic network; chemical synthesis; Dendrobium catenatum; Hydroxylation; Intermediates; ligands; Medicinal plants; Methylation; methyltransferases; O‐methyltransferase; Piceatannol; polymethoxylated bibenzyl; regioselectivity; species; Species diversity; Substrate preferences; Substrates; chemical synthesis; O‐methyltransferase; Dendrobium catenatum; biosynthetic network; polymethoxylated bibenzyl
• ISSN: 0960-7412; 1365-313X; 1365-313X
• DOI: 10.1111/tpj.16962

SUMMARY Erianin, crepidatin, and chrysotobibenzyl are typical medicinal polymethoxylated bibenzyls (PMBs) that are commercially produced in Dendrobium species. PMBs' chemo‐diversity is mediated by the manifold combinations of O‐methylation and hydroxylation in a definite order, which remains unsolved. To unequivocally elucidate the methylation mechanism of PMBs, 15 possible intermediates in the biosynthetic pathway of PMBs were chemically synthesized. DcOMT1–5 were highly expressed in tissues where PMBs were biosynthesized, and their expression patterns were well‐correlated with the accumulation profiles of PMBs. Moreover, cell‐free orthogonal tests based on the synthesized intermediates further confirmed that DcOMT1‐5 exhibited distinct substrate preferences and displayed hydroxyl‐group regiospecificity during the sequential methylation process. The stepwise methylation of PMBs was discovered from SAM to dihydro‐piceatannol (P) in the following order: P → 3‐MeP → 4‐OH‐3‐MeP → 4‐OH‐3,5‐diMeP → 3,3'(4′),5‐triMeP → 3,4,4′,5‐tetraMeP (erianin) or 3,3′,4,5‐tetraMeP (crepidatin) → 3,3′,4,4′,5‐pentaMeP (chrysotobibenzyl). Furthermore, the regioselectivities of DcOMTs were investigated by ligand docking analyses which corresponded precisely with the catalytic activities. In summary, the findings shed light on the sequential catalytic mechanisms of PMB biosynthesis and provide a comprehensive PMB biosynthetic network in D. catenatum. The knowledge gained from this study may also contribute to the development of plant‐based medicinal applications and the production of high‐value PMBs. Significance Statement Our study is an icebreaker for the dissection of the PMBs biosynthesis pathway. By chemically synthesising intermediates, substrate docking, and enzyme activity assay, we ultimately summarised the catalytic laws of PMBs in D. catenatum. Molecular docking provided an in‐depth understanding of the structural framework for the substrate binding and transmethylation activities of DcOMTs, thereby substantially increasing the understanding of its distinctive substrate specificities, regioselectivities, and catalytic efficiencies, elevating evidence for bioengineering of the high‐value and novel O‐methylated bibenzyls products.