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 composition comprising an engineered non-plant cell that contains a polynucleotide encoding a polypeptide having thebaine synthase activity that is at least 70% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
2. The composition of claim 1 , wherein the polynucleotide encodes a polypeptide that is at least 80% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
3. The composition of claim 1 , wherein the polynucleotide encodes a polypeptide that is at least 85% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
4. The composition of claim 1 , wherein the polynucleotide encodes a polypeptide that is at least 90% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
5. The composition of claim 1 , wherein the polynucleotide encodes a polypeptide that is at least 95% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
6. The composition of claim 1 , wherein the polynucleotide encodes a polypeptide 100% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.
7. The composition of claim 1 , wherein the non-plant cell is a yeast cell.
8. The composition of claim 7 , wherein the yeast cell is a Saccharomyces cerevisiae cell or a Yarrowia lipolytica cell.
9. The composition of claim 1 , wherein the polynucleotide is integrated into an expression vector.
10. An expression vector comprising a polynucleotide encoding a polypeptide having thebaine synthase activity that is at least 70% identical to the amino acid sequence of SEQ ID NO: 29 or SEQ ID NO:900.
11. The expression vector of claim 10 , wherein the expression vector further comprises polynucleotide elements capable of controlling expression of the polynucleotide in a cell.
12. The expression vector of claim 11 , wherein the polynucleotide elements capable of controlling expression in a host cell comprises a promoter, a transcriptional terminator, a transcriptional enhancer, or combinations thereof.
13. The expression vector of claim 10 , further comprising one or more polynucleotide elements encoding marker genes, one or more origins of replication, genetic elements required for the integration of the vector or a portion thereof into a host cell's genome, or combinations thereof.
14. The expression vector of claim 10 , wherein the polynucleotide encodes a polypeptide that is at least 80% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO: 900.
15. The expression vector of claim 10 , wherein the polynucleotide encodes a polypeptide that is at least 85% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO: 900.
16. The expression vector of claim 10 , wherein the polynucleotide encodes a polypeptide that is at least 90% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO: 900.
17. The expression vector of claim 10 , wherein the polynucleotide encodes a polypeptide that is at least 95% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO: 900.
18. The expression vector of claim 10 , wherein the polynucleotide encodes a polypeptide 100% identical to the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:900.