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Frusciante S, Demurtas OC, Sulli M, et al. Heterologous expression of Bixa orellana cleavage dioxygenase 4-3 drives crocin but not bixin biosynthesis. Plant Physiol. 2021;doi: 10.1093/plphys/kiab583.
Frusciante, S., Demurtas, O. C., Sulli, M., Mini, P., Aprea, G., Diretto, G., Karcher, D., Bock, R., & Giuliano, G. (2021). Heterologous expression of Bixa orellana cleavage dioxygenase 4-3 drives crocin but not bixin biosynthesis. Plant physiology, . https://doi.org/10.1093/plphys/kiab583
Frusciante, Sarah, et al. "Heterologous expression of Bixa orellana cleavage dioxygenase 4-3 drives crocin but not bixin biosynthesis." Plant physiology vol. (2021). doi: https://doi.org/10.1093/plphys/kiab583
Frusciante S, Demurtas OC, Sulli M, Mini P, Aprea G, Diretto G, Karcher D, Bock R, Giuliano G. Heterologous expression of Bixa orellana cleavage dioxygenase 4-3 drives crocin but not bixin biosynthesis. Plant Physiol. 2021 Dec 17; doi: 10.1093/plphys/kiab583. Epub 2021 Dec 17. PMID: 34919714.
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Plant Physiol. 2021 Dec 17; doi: 10.1093/plphys/kiab583. Epub 2021 Dec 17.

Heterologous expression of Bixa orellana cleavage dioxygenase 4-3 drives crocin but not bixin biosynthesis.

Plant physiology

Sarah Frusciante, Olivia Costantina Demurtas, Maria Sulli, Paola Mini, Giuseppe Aprea, Gianfranco Diretto, Daniel Karcher, Ralph Bock, Giovanni Giuliano

Affiliations

  1. Italian national Agency for New technologies, Energy, and Sustainable Development (ENEA), Casaccia Research Center, 00123 Roma, Italy.
  2. Max-Planck-Institut für Molekulare Pflanzenphysiologie, Am Mühlenberg 1, 14476 Potsdam-Golm, Germany.

PMID: 34919714 DOI: 10.1093/plphys/kiab583

Abstract

Annatto (Bixa orellana) is a perennial shrub native to the Americas, and bixin, derived from its seeds, is a methoxylated apocarotenoid used as a food and cosmetic colorant. Two previous reports claimed to have isolated the carotenoid cleavage dioxygenase (CCD) responsible for the production of the putative precursor of bixin, the C24 apocarotenal bixin dialdehyde. We re-assessed the activity of six Bixa CCDs and found that none of them produced substantial amounts of bixin dialdehyde in Escherichia coli. Unexpectedly, BoCCD4-3 cleaved different carotenoids (lycopene, β-carotene and zeaxanthin) to yield the C20 apocarotenal crocetin dialdehyde, the known precursor of crocins, which are glycosylated apocarotenoids accumulated in saffron stigmas. BoCCD4-3 lacks a recognizable transit peptide but localized to plastids, the main site of carotenoid accumulation in plant cells. Expression of BoCCD4-3 in Nicotiana benthamiana leaves (transient expression), tobacco (Nicotiana tabacum) leaves (chloroplast transformation, under the control of a synthetic riboswitch) and, in conjunction with a saffron crocetin glycosyl transferase, in tomato (Solanum lycopersicum) fruits (nuclear transformation) led to high levels of crocin accumulation, reaching the highest levels (>100 µg/g dry weight) in tomato fruits, which also showed a crocin profile similar to that found in saffron, with highly glycosylated crocins as major compounds. Thus, while the bixin biosynthesis pathway remains unresolved, BoCCD4-3 can be used for the metabolic engineering of crocins in a wide range of different plant tissues.

© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.

Keywords: Bixa orellana ; Crocus sativus ; Nicotiana ; annatto; carotenoid cleavage dioxygenase; crocins; metabolic engineering; tomato

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