Discovery and engineering of colchicine alkaloid biosynthesis

Discovery of a near-complete colchicine biosynthetic pathway enables the engineered production of the tropolone-containing alkaloidN-formyldemecolcine from amino acid precursors inNicotiana benthamiana. Few complete pathways have been established for the biosynthesis of medicinal compounds from plants. Accordingly, many plant-derived therapeutics are isolated directly from medicinal plants or plant cell culture(1). A lead example is colchicine, a US Food and Drug Administration (FDA)-approved treatment for inflammatory disorders that is sourced fromColchicumandGloriosaspecies(2-5). Here we use a combination of transcriptomics, metabolic logic and pathway reconstitution to elucidate a near-complete biosynthetic pathway to colchicine without prior knowledge of biosynthetic genes, a sequenced genome or genetic tools in the native host. We uncovered eight genes fromGloriosa superbafor the biosynthesis ofN-formyldemecolcine, a colchicine precursor that contains the characteristic tropolone ring and pharmacophore of colchicine(6). Notably, we identified a non-canonical cytochrome P450 that catalyses the remarkable ring expansion reaction that is required to produce the distinct carbon scaffold of colchicine. We further used the newly identified genes to engineer a biosynthetic pathway (comprising 16 enzymes in total) toN-formyldemecolcine inNicotiana benthamianastarting from the amino acids phenylalanine and tyrosine. This study establishes a metabolic route to tropolone-containing colchicine alkaloids and provides insights into the unique chemistry that plants use to generate complex, bioactive metabolites from simple amino acids.