RESUMO
In Coptis japonica cell cultures an alternative pathway has been discovered which leads from (S)-tetrahydrocolumbamine via (S)-canadine to berberine. The two enzymes involved have been partially purified. (S)-Tetrahydrocolumbamine is stereospecifically transformed into (S)-canadine under formation of the methylenedioxy bridge in ring A. This new enzyme was named (S)-canadine synthase. (S)-Canadine in turn is stereospecifically dehydrogenated to berberine by an oxidase, (S)-canadine oxidase (COX), which was partially purified (25-fold). This enzyme has many physical properties in common with the already known (S)-tetrahydroprotoberberine oxidase from Berberis but grossly differs from the latter enzyme in its cofactor requirement (Fe) and its substrate specificity. Neither (S)-norreticuline nor (S)-scoulerine serves as substrate for the Coptis enzyme, while both substrates are readily oxidized by the Berberis enzyme. The four terminal enzymes catalyzing the pathway from (S)-reticuline to berberine are housed in Berberis as well as in Coptis in smooth vesicles with a density of ρ=1.14 g/ml. These vesicles have been enriched and characterized by electron microscopy.
