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J Exp Bot. 2017 Jul 20;68(16):4663-4677. doi: 10.1093/jxb/erx277.

Evolutionarily distinct carotenoid cleavage dioxygenases are responsible for crocetin production in Buddleja davidii.

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Instituto Botánico, Departamento de Ciencia y Tecnología Agroforestal y Genética, Facultad de Farmacia, Universidad de Castilla-La Mancha, Campus Universitario s/n, 02071 Albacete, Spain.
Facultad de Ciencias Ambientales y Bioquímica, Universidad de Castilla-La Mancha, Campus Tecnológico de la Fábrica de Armas, Avda, Carlos III s/n, E-45071 Toledo, Spain.
Italian National Agency for New Technologies, Energy, and Sustainable Development, Casaccia Research Centre, 00123 Rome, Italy.
Instituto de Biología Molecular y Celular de Plantas, Consejo Superior de Investigaciones Científicas, Universidad Politécnica de Valencia, Camino de Vera s/n, 46022 Valencia, Spain.


Crocetin, one of the few colored apocarotenoids known in nature, is present in flowers and fruits and has long been used medicinally and as a colorant. Saffron is the main source of crocetin, although a few other plants produce lower amounts of this apocarotenoid. Notably, Buddleja davidii accumulates crocetin in its flowers. Recently, a carotenoid dioxygenase cleavage enzyme, CCD2, has been characterized as responsible for crocetin production in Crocus species. We searched for CCD2 homologues in B. davidii and identified several CCD enzymes from the CCD1 and CCD4 subfamilies. Unexpectedly, two out of the three CCD4 enzymes, namely BdCCD4.1 and BdCCD4.3, showed 7,8;7',8' activity in vitro and in vivo over zeaxanthin. In silico analyses of these enzymes and CCD2 allowed the determination of key residues for this activity. Both BdCCD4 genes are highly expressed during flower development and transcripts levels parallel the accumulation of crocins in the petals. Phylogenetic analysis showed that BdCCD4.2 grouped with almost all the characterized CCD4 enzymes, while BdCCD4.1 and BdCCD4.3 form a new sub-cluster together with CCD4 enzymes from certain Lamiales species. The present study indicates that convergent evolution led to the acquisition of 7,8;7',8' apocarotenoid cleavage activity in two separate CCD enzyme families.


Carotenoid dioxygenase cleavage; Lamiales; crocetin; flowers; zeaxanthin

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