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Engineering Saccharomyces cerevisiae for the Overproduction of ?-Ionone and Its Precursor ?-Carotene.

ABSTRACT: ?-ionone is a commercially attractive industrial fragrance produced naturally from the cleavage of the pigment ?-carotene in plants. While the production of this ionone is typically performed using chemical synthesis, environmentally friendly and consumer-oriented biotechnological production is gaining increasing attention. A convenient cell factory to address this demand is the yeast Saccharomyces cerevisiae. However, current ?-ionone titers and yields are insufficient for commercial bioproduction. In this work, we optimized S. cerevisiae for the accumulation of high amounts of ?-carotene and its subsequent conversion to ?-ionone. For this task, we integrated systematically the heterologous carotenogenic genes (CrtE, CrtYB and CrtI) from Xanthophyllomyces dendrorhous using markerless genome editing CRISPR/Cas9 technology; and evaluated the transcriptional unit architecture (bidirectional or tandem), integration site, and impact of gene dosage, first on ?-carotene accumulation, and later, on ?-ionone production. A single-copy insertion of the carotenogenic genes in high expression loci of the wild-type yeast CEN.Pk2 strain yielded 4 mg/gDCW of total carotenoids, regardless of the transcriptional unit architecture employed. Subsequent fine-tuning of the carotenogenic gene expression enabled reaching 16 mg/gDCW of total carotenoids, which was further increased to 32 mg/gDCW by alleviating the known pathway bottleneck catalyzed by the hydroxymethylglutaryl-CoA reductase (HMGR1). The latter yield represents the highest total carotenoid concentration reported to date in S. cerevisiae for a constitutive expression system. For ?-ionone synthesis, single and multiple copies of the carotene cleavage dioxygenase 1 (CCD1) gene from Petunia hybrida (PhCCD1) fused with a membrane destination peptide were expressed in the highest ?-carotene-producing strains, reaching up to 33 mg/L of ?-ionone in the culture medium after 72-h cultivation in shake flasks. Finally, interrogation of a contextualized genome-scale metabolic model of the producer strains pointed to PhCCD1 unspecific cleavage activity as a potentially limiting factor reducing ?-ionone production. Overall, the results of this work constitute a step toward the industrial production of this ionone and, more broadly, they demonstrate that biotechnological production of apocarotenoids is technically feasible.

SUBMITTER: Lopez J

PROVIDER: S-EPMC7556307 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Engineering <i>Saccharomyces cerevisiae</i> for the Overproduction of β-Ionone and Its Precursor β-Carotene.

López Javiera J Bustos Diego D Camilo Conrado C Arenas Natalia N Saa Pedro A PA Agosin Eduardo E

Frontiers in bioengineering and biotechnology 20200930

β-ionone is a commercially attractive industrial fragrance produced naturally from the cleavage of the pigment β-carotene in plants. While the production of this ionone is typically performed using chemical synthesis, environmentally friendly and consumer-oriented biotechnological production is gaining increasing attention. A convenient cell factory to address this demand is the yeast <i>Saccharomyces cerevisiae</i>. However, current β-ionone titers and yields are insufficient for commercial bio ...[more]

PMID: 33102463

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Dataset Information

Engineering Saccharomyces cerevisiae for the Overproduction of ?-Ionone and Its Precursor ?-Carotene.

Publications

Engineering <i>Saccharomyces cerevisiae</i> for the Overproduction of β-Ionone and Its Precursor β-Carotene.

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