Publications

• Heterodimeric O-methyltransferases involved in the biosynthesis of noscapine in opium poppy.

  Park M.R., Chen X., Lang D.E., Ng K.K.S., Facchini P.J.

  Noscapine biosynthesis in opium poppy involves three characterized O-methyltransferases (OMTs) and a fourth responsible for the 4'-methoxyl on the phthalide isoquinoline scaffold. The first three enzymes are homodimers, whereas the latter is a heterodimer encoded by two linked genes (OMT2 and OMT3 ... >> More

  Noscapine biosynthesis in opium poppy involves three characterized O-methyltransferases (OMTs) and a fourth responsible for the 4'-methoxyl on the phthalide isoquinoline scaffold. The first three enzymes are homodimers, whereas the latter is a heterodimer encoded by two linked genes (OMT2 and OMT3). Neither OMT2 nor OMT3 form stable homodimers, but yield a substrate-specific heterodimer when their genes are co-expressed in Escherichia coli. The only substrate, 4'-O-desmethyl-3-O-acetylpapaveroxine, is a seco-berbine pathway intermediate that undergoes ester hydrolysis subsequent to 4'-O-methylation leading to the formation of narcotine hemiacetal. In the absence of 4'-O-methylation, a parallel pathway yields narcotoline hemiacetal. Dehydrogenation produces noscapine and narcotoline from the corresponding hemiacetals. Phthalide isoquinoline intermediates with a 4'-hydroxyl (i.e. narcotoline and narcotoline hemiacetal), or the corresponding 1-hydroxyl on protoberberine intermediates, were not accepted. Norcoclaurine 6OMT, which shares 81% amino acid sequence identity with OMT3, also formed a functionally similar heterodimer with OMT2. Suppression of OMT2 transcript levels in opium poppy increased narcotoline accumulation, whereas reduced OMT3 transcript abundance caused no detectable change in the alkaloid phenotype. Opium poppy chemotype Marianne accumulates high levels of narcotoline and showed no detectable OMT2:OMT3 activity. Compared with the active subunit from the Bea's Choice chemotype, Marianne OMT2 exhibited a single S122Y mutation in the dimerization domain that precluded heterodimer formation based on homology models. Both subunits contributed to the formation of the substrate-binding domain, although site-directed mutagenesis revealed OMT2 as the active subunit. The occurrence of physiologically relevant OMT heterodimers increases the catalytic diversity of enzymes derived from a smaller number of gene products. << Less

  Plant J. 95:252-267(2018) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• Engineering biosynthesis of the anticancer alkaloid noscapine in yeast.

  Li Y., Smolke C.D.

  Noscapine is a potential anticancer drug isolated from the opium poppy Papaver somniferum, and genes encoding enzymes responsible for the synthesis of noscapine have been recently discovered to be clustered on the genome of P. somniferum. Here, we reconstitute the noscapine gene cluster in Sacchar ... >> More

  Noscapine is a potential anticancer drug isolated from the opium poppy Papaver somniferum, and genes encoding enzymes responsible for the synthesis of noscapine have been recently discovered to be clustered on the genome of P. somniferum. Here, we reconstitute the noscapine gene cluster in Saccharomyces cerevisiae to achieve the microbial production of noscapine and related pathway intermediates, complementing and extending previous in planta and in vitro investigations. Our work provides structural validation of the secoberberine intermediates and the description of the narcotoline-4'-O-methyltransferase, suggesting this activity is catalysed by a unique heterodimer. We also reconstitute a 14-step biosynthetic pathway of noscapine from the simple alkaloid norlaudanosoline by engineering a yeast strain expressing 16 heterologous plant enzymes, achieving reconstitution of a complex plant pathway in a microbial host. Other engineered yeasts produce previously inaccessible pathway intermediates and a novel derivative, thereby advancing protoberberine and noscapine related drug discovery. << Less

  Nat. Commun. 7:12137-12137(2016) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.