Heterologous expression of alkaloid biosynthetic genes--a review.
Tetrahydrobenzylisoquinoline alkaloids comprise a diverse class of secondary metabolites with many pharmacologically active members. The biosynthesis at the enzyme level of at-least two tetrahydrobenzylisoquinoline alkaloids, the benzophenanthridine alkaloid sanguinarine in the California poppy, Eschscholtzia californica, and the bisbenzylisoquinoline alkaloid berbamunine in barberry, Berberis stolonifera, has been elucidated in detail starting from the aromatic amino acid (aa) L-tyrosine. In an initial attempt to develop alternate systems for the production of medicinally important alkaloids, one enzyme from each pathway (BBE, a covalently flavinylated enzyme of benzophenanthridine alkaloid biosynthesis and CYP80, a phenol coupling cytochrome P-450-dependent oxidase of bisbenzylisoquinoline alkaloid biosynthesis) has been purified to homogeneity, a partial aa sequence determined, and the corresponding cDNAs isolated with aid of synthetic oligos based on the aa sequences. The recombinant enzymes were actively expressed in Spotloptera frugiperda Sf9 cells using a baculovirus vector, purified and then characterized. Insect cell culture has proven to be a powerful system for the overexpression of alkaloid biosynthetic genes.
Kutchan
T M
TM
Laboratorium für Molekulare Biologie, Universität München, Germany. u61116@sunmail.lrz-muenchen.de
D000429:Alcohol Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D000470:Alkaloids / Q000096:biosynthesis*; D053119:Benzophenanthridines; D044182:Benzylisoquinolines; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000235:genetics; D015870:Gene Expression; D017343:Genes, Plant; D007546:Isoquinolines; D010089:Oxidoreductases, N-Demethylating / Q000096:biosynthesis* / Q000235:genetics; D010940:Plant Proteins / Q000096:biosynthesis* / Q000235:genetics; D010944:Plants / Q000201:enzymology / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis
D000470:Alkaloids; D053119:Benzophenanthridines; D044182:Benzylisoquinolines; D007546:Isoquinolines; D010940:Plant Proteins; D011994:Recombinant Proteins; C102663:berbamunine; C092555:cytochrome P-450 CYP80A1 (Berberis stolonifera); D003577:Cytochrome P-450 Enzyme System; C005705:sanguinarine; D000429:Alcohol Oxidoreductases; C071690:reticuline oxidase; D010089:Oxidoreductases, N-Demethylating