Patent ID: 9322039
Filing Date: 2016-04-26
CPC Classification: C12N,C12P

Claim Text:
1. A method of preparing a metabolite of tyrosine that is a benzylisoquinoline alkaloid product, the method comprising: a) culturing an engineered non-plant cell under conditions suitable for protein production, said engineered non-plant cell comprising three heterologous coding sequences, wherein the three heterologous coding sequences encode a first, second, and third enzyme, respectively, that are involved in a metabolic pathway that converts the tyrosine into the benzylisoquinoline alkaloid product, wherein the first, second, and third enzymes are operably connected along the metabolic pathway; b) optionally adding tyrosine to the cell culture; and c) recovering the benzylisoquinoline alkaloid product from the cell culture, wherein the benzylisoquinoline alkaloid product is selected from the group consisting of a norcoclaurine, coclaurine, N-methylcoclaurine, 3′-hydroxy-N-methylcoclaurine, reticuline, 6-O-methyl-norlaudanosoline, 6-O-methyl-laudanosoline, laudanine, scoulerine, tetrahydrocolumbamine, canadine, salutaridine, salutaridinol, salutaridinol-7-O-acetate, and thebaine, and wherein each of the first, second, and third enzymes involved in the metabolic pathway that produces the benzylisoquinoline alkaloid product is selected from the group consisting of L-tyrosine/dopa decarboxylase 1, L-tyrosine/dopa decarboxylase 2, Cytochrome P450 2D6, NADPH p450 reductase, Polyphenyloxidase, Tyrosine hydroxylase, GTPcyclohydrolase I, Monoamine oxidase A, Tyramine oxidase, Aromatic amino acid transaminase, Phenylpyruvate decarboxylase, Norcoclaurine synthase, Norcoclaurine 6-O-methyltransferase, Coclaurine-N-methyltransferase, Cytochrome P450 80B1, 4-O-methyltransferase, Berberine bridge enzyme, Reticuline 7-O-methyltransferase, Scoulerine 9-O-methyltransferase, Canadine synthase, Salutaridine reductase, Salutaridinol 7-O-acetyltransferase, Codeine reductase, and Berbamunine synthase.