EC Number   |
|---|
    1.1.1.248 | cloned into the pMyr vector and expressed as fusion protein with a myristylation sequence, which anchors the target protein to the yeast membrane. Fusion proteins (pSOS/SalAT and pMyr/SalR) coexpressed in the Saccharomyces cerevisiae cdc25H strain. SalR amplified from the recombinant plasmid SalR/pQE-30. The 0.9-kb DNA fragment ligated into an NheI/EcoRI-digested pET28a expression vector. SalR/pET28a expression construct transformed into Escherichia coli BL21(DE3)RIL |
| 1.1.1.248 | expression in Escherichia coli |
| 1.1.1.248 | gene salR, DNA and amino acid sequence determination and analysis, expression profile, overexpression in Escherichia coli strain SG13009 as His6-tagged enzyme |
| 1.1.1.248 | gene salR, expression of N-terminally His-tagged enzyme in Escherichia coli, a selenomethionine-substituted SalR is produced by inhibition of the methionine biosynthetic pathway with the same expression vector and Escherichia coli strain used for expression of wild-type SalR |
| 1.1.1.248 | heterologously overexpressed in its active form from Papaver somniferum |
| 1.1.1.248 | overexpression in Escherichia coli SG13009 |
| 1.1.1.248 | real-time quantitative PCR expression analysis |
| 1.1.1.248 | salR, DNA and amino acid sequence determination and analysis, overexpression of the His-tagged enzyme in Escherichia coli |
| 1.1.1.248 | salutaridine synthase (PsSAS), salutaridine reductase (PsSAR) and salutaridinol acetyltransferase (PsSAT) are functionally coexpressed in Saccharomyces cerevisiae and optimization of the pH conditions allowed for productive spontaneous rearrangement of salutaridinol-7-O-acetate and synthesis of thebaine from (R)-reticuline. A 7-gene pathway for the production of codeine and morphine from (R)-reticuline is reconstituted. Yeast cell feeding assays using (R)-reticuline, salutaridine or codeine as substrates show that all enzymes are functionally coexpressed 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 Saccharomyces cerevisiae and pave the way for their complete synthesis in recombinant microbes |
