Polynucleotides and polypeptides useful for making alkaloid compounds
Polynucleotides and polypeptides useful in the manufacture of a class of chemical compounds known as alkaloids are provided. The polynucleotides and polypeptides may be used to synthesize alkaloids, including reticuline, thebaine and morphine, in vivo and in vitro. The polynucleotides further may be used to examine the presence of the polynucleotides in a cell or a cell extract, and to modulate expression thereof in living cells.
1. A yeast cell comprising a first polynucleotide encoding a polypeptide having thebaine synthase activity comprising an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: SEQ ID NO: 29 or SEQ ID NO: 900.
2. The yeast cell of claim 1 , further comprising a second polynucleotide having purine permease activity encoding a polypeptide comprising an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: SEQ ID NO: 779 or SEQ ID NO: 901.
3. The yeast cell of claim 2 , further comprising a third polynucleotide that links the first and second polynucleotides.
4. The yeast cell of claim 2 , wherein the first and second polynucleotides are integrated, respectively, into first and second expression vectors.
5. The yeast cell of claim 1 , wherein the cell is a Saccharomyces cerevisiae cell or a Yarrowia lipolytica cell.
6. A method of reducing the expression of a polynucleotide encoding a polypeptide having thebaine synthase activity comprising an amino acid sequence that is at least 90% identical to an amino acid sequence selected from the group consisting of: SEQ ID NO: 29 or SEQ ID NO: 900 in a cell expressing the polynucleotide, the method comprising silencing the expression of the polynucleotide in the cell,
wherein the cell is a Papaver somniferum cell or the cell of claim 1 .
7. The yeast cell of claim 1 , wherein the first polynucleotide encodes a polypeptide comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 29 or SEQ ID NO: 900.
8. The yeast cell of claim 2 , wherein the second polynucleotide encodes a polypeptide comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 779 or SEQ ID NO: 901.
9. The yeast cell of claim 1 , wherein the first polynucleotide encodes a polypeptide comprising an amino acid sequences that is at least 97% identical to SEQ ID NO: 29 or SEQ ID NO: 900.
10. The yeast cell of claim 2 , wherein the second polynucleotide encodes a polypeptide comprising an amino acid sequence that is at least 97% identical to SEQ ID NO: 779 or SEQ ID NO: 901.
11. The yeast cell of claim 1 , wherein the first polynucleotide comprises a nucleic acid sequence capable of hybridizing under at least moderately stringent hybridization conditions with the nucleic acid sequence of SEQ ID NO: 6 or SEQ ID NO: 372.