Major anthocyanin biosynthesis in the brilliant crimson petals from Erythrina crista-galli L

•We identified a major anthocyanin in Erythrina crista-galli petals.•We reported the first biochemical characterization of 3GGT from woody plant.•The Ec3GGT activity was inhibited by divalent cations and SH-blocking reagents.•Both anthocyanin and flavonol were preferred substrates of Ec3GGT.•The molecular mass of Ec3GGT was estimated to be larger than known UFGTs.

The anthocyanins in the brilliant crimson petals from Erythrina crista-galli L. were extracted with 10% AcOH, and the extract was purified by Amberlite XAD-7 column chromatography, paper chromatography, and preparative HPLC. The structure of a major anthocyanin was confirmed as cyanidin 3-O-sophoroside by 1H and 13C NMR and FABMS. Then, the catalysis of UDP-glucose: flavonoid 3-O-glucoside glucosyltransferase (3GGT) was predicted at the final step in the anthocyanin biosynthesis. In this study, the 3GGT was purified approximately to 800-fold from E. crista-galli petals by eight-steps purification procedures. The purified enzyme (Ec3GGT) catalyzed only the 2″-O-glucosylation of flavonoid 3-O-glucoside to produce flavonoid 3-O-sophoroside. Ec3GGT showed a pH optimum around 7.5. The molecular weight and isoelectric point were estimated to be about 82 kDa and 6.5, respectively; the molecular weight was larger than known UDP-sugar: flavonoid glycosyltransferases (UFGTs). The enzyme did not require metal cofactors but inhibited by Zn2+, Co2+ and Cu2+, and it activity was inhibited by SH-blocking reagents such as NEM and PCMB. Both anthocyanidin 3-O-glucosides and flavonol 3-O-glucosides were favorable acceptor substrates of the enzyme, whereas no activity was shown toward C-glucosylflavones such as isovitexin, which is a minor component in the petals. These results showed that Ec3GGT involved in the last step of biosynthesis of a major anthocyanin in petals.

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