Synthesis of Morphinan Alkaloids in Saccharomyces cerevisiae

PLoS One. 2015 Apr 23;10(4):e0124459. doi: 10.1371/journal.pone.0124459. eCollection 2015.

Abstract

Morphinan alkaloids are the most powerful narcotic analgesics currently used to treat moderate to severe and chronic pain. The feasibility of morphinan synthesis in recombinant Saccharomyces cerevisiae starting from the precursor (R,S)-norlaudanosoline was investigated. Chiral analysis of the reticuline produced by the expression of opium poppy methyltransferases showed strict enantioselectivity for (S)-reticuline starting from (R,S)-norlaudanosoline. In addition, the P. somniferum enzymes salutaridine synthase (PsSAS), salutaridine reductase (PsSAR) and salutaridinol acetyltransferase (PsSAT) were functionally co-expressed in S. cerevisiae and optimization of the pH conditions allowed for productive spontaneous rearrangement of salutaridinol-7-O-acetate and synthesis of thebaine from (R)-reticuline. Finally, we reconstituted a 7-gene pathway for the production of codeine and morphine from (R)-reticuline. Yeast cell feeding assays using (R)-reticuline, salutaridine or codeine as substrates showed that all enzymes were functionally co-expressed in yeast and that activity of salutaridine reductase and codeine-O-demethylase likely limit flux to morphine synthesis. The results of this study describe a significant advance for the synthesis of morphinans in S. cerevisiae and pave the way for their complete synthesis in recombinant microbes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, High Pressure Liquid
  • Fourier Analysis
  • Mass Spectrometry
  • Morphinans / chemistry
  • Morphinans / metabolism*
  • Saccharomyces cerevisiae / metabolism*
  • Stereoisomerism

Substances

  • Morphinans

Grants and funding

This work was financially supported by Genome Canada (http://www.genomecanada.ca) and Genome Québec (http://www.genomequebec.com). VJJM is supported by a Canada Research Chair (http://www.chairs-chaires.gc.ca). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.