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

López, Javiera; Bustos, D.P. Jaramillo; Camilo, Conrado; Arenas, Natalia; Saa, Pedro A.; Agosín, Eduardo

doi:10.3389/fbioe.2020.578793

Cited by 32 publications

(23 citation statements)

References 43 publications

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“…However, the small number of selection markers, integration sites, and genetic arrangements impedes strain design, as the acceptable gene dosage (copy number) for construction is restricted. CRISPR/Cas9 technology advancement in lees minimizes the need for selection markers, particularly when the transformation of S. cerevisiae has proven to be highly successful [94,95].…”

Section: Microbial Production and Biotransformation Of β-Iononementioning

confidence: 99%

“…Instead, using the oleaginous yeast Yarrowia lipolytica, the authors of [90] obtained titers of 60 mg/L in shake flasks and 380 mg/L of β-ionone in a fermenter. More recently, López et al [95] developed several recombinant S. cerevisiae strains capable of producing increasing amounts of β-ionone (33 mg/L) and its precursor, β-carotene (33 mg/L), in the culture medium after 72 h cultivation in shake flasks.…”

Section: Microbial Production and Biotransformation Of β-Iononementioning

confidence: 99%

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β-Ionone: Its Occurrence and Biological Function and Metabolic Engineering

Paparella

Shaltiel-Harpaza

Ibdah

2021

Plants

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β-Ionone is a natural plant volatile compound, and it is the 9,10 and 9’,10’ cleavage product of β-carotene by the carotenoid cleavage dioxygenase. β-Ionone is widely distributed in flowers, fruits, and vegetables. β-Ionone and other apocarotenoids comprise flavors, aromas, pigments, growth regulators, and defense compounds; serve as ecological cues; have roles as insect attractants or repellants, and have antibacterial and fungicidal properties. In recent years, β-ionone has also received increased attention from the biomedical community for its potential as an anticancer treatment and for other human health benefits. However, β-ionone is typically produced at relatively low levels in plants. Thus, expressing plant biosynthetic pathway genes in microbial hosts and engineering the metabolic pathway/host to increase metabolite production is an appealing alternative. In the present review, we discuss β-ionone occurrence, the biological activities of β-ionone, emphasizing insect attractant/repellant activities, and the current strategies and achievements used to reconstruct enzyme pathways in microorganisms in an effort to to attain higher amounts of the desired β-ionone.

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Section: Microbial Production and Biotransformation Of β-Iononementioning

confidence: 99%

Section: Microbial Production and Biotransformation Of β-Iononementioning

confidence: 99%

β-Ionone: Its Occurrence and Biological Function and Metabolic Engineering

Paparella

Shaltiel-Harpaza

Ibdah

2021

Plants

View full text Add to dashboard Cite

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“…The synthesis of β-ionone catalyzed using synthetic biology and biological enzymes has been reported, but not the synthesis of 3-hydroxy-β-ionone [9,[39][40][41]. The effects of enzyme and substrate concentrations on the biocatalytic process of EhLCD are important owing to its practical applications.…”

Section: Production Of 3-hydroxy-β-ionone From Lutein By Ehlcdmentioning

confidence: 99%

Characterization of a Novel Lutein Cleavage Dioxygenase, EhLCD, from Enterobacter hormaechei YT-3 for the Enzymatic Synthesis of 3-Hydroxy-β-ionone from Lutein

et al. 2021

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3-Hydroxy-β-ionone, a flavor and fragrance compound with fruity violet-like characteristics, is widely applied in foodstuff and beverages, and is currently produced using synthetic chemistry. In this study, a novel lutein cleavage enzyme (EhLCD) was purified and characterized from Enterobacter hormaechei YT-3 to convert lutein to 3-hydroxy-β-ionone. Enzyme EhLCD was purified to homogeneity by ammonium sulfate precipitation, Q-Sepharose, phenyl-Sepharose, and Superdex 200 chromatography. The molecular mass of purified EhLCD, obtained by SDS-PAGE, was approximately 50 kDa. The enzyme exhibited the highest activity toward lutein, followed by zeaxanthin, β-cryptoxanthin, and β-carotene, suggesting that EhLCD exhibited higher catalytic efficiency for carotenoid substrates bearing 3-hydroxy-ionone rings. Isotope-labeling experiments showed that EhLCD incorporated oxygen from O2 into 3-hydroxy-β-ionone and followed a dioxygenase reaction mechanism for different carotenoid substrates. These results indicated that EhLCD is the first characterized bacterial lutein cleavage dioxygenase. Active EhLCD was also confirmed to be a Fe2+-dependent protein with 1 molar equivalent of non-haem Fe2+. The purified enzyme displayed optimal activity at 45 °C and pH 8.0. The optimum concentrations of the substrate, enzyme, and Tween 40 for 3-hydroxy-β-ionone production were 60 mM lutein/L, 1.5 U/mL, and 2% (w/v), respectively. Under optimum conditions, EhLCD produced 3-hydroxy-β-ionone (637.2 mg/L) in 60 min with a conversion of 87.0% (w/w), indicating that this enzyme is a potential candidate for the enzymatic synthesis of 3-hydroxy-β-ionone in biotechnological applications.

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“…In addition, β-ionone production in other microorganisms has also been attempted. For example, in S. cerevisiae, titers of 33 mg/L and 184 mg/L in shake flasks have been reported; [10,11] while in the oleaginous yeast Yarrowia lipolytica, its yield reached up to 358 mg/L in shake-flask fermentation and 1 g/L in fed-batch fermentation, the highest β-ionone titer of engineered strain reported to date. [12] Nevertheless, the major problem restricting the efficient synthesis of β-ionone is that the catalytic activity of CCDs in the synthetic pathway is low, and specificity is not strong leads to the low production of β-ionone.…”

Section: Introductionmentioning

confidence: 99%

Overexpression and Characterization of a Novel Plant Carotenoid Cleavage Dioxygenase 1 from Morus notabilis

Fan

Zhu

et al. 2021

Chemistry & Biodiversity

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Synthesis of β-ionone in microbial cell factories is limited by the efficiency of carotenoid cleavage dioxygenases (CCDs). To obtain genes responsible for specific cleavage of carotenoids generating β-ionone, a novel carotenoid cleavage dioxygenase 1 from Morus notabilis was cloned and overexpressed in Escherichia coli. The MnCCD1 protein was able to cleave a variety of carotenoids at the positions 9, 10 (9', 10') to produce βionone, 3-hydroxy-4-oxo-β-ionone, 3-hydroxy-β-ionone, and 3-hydroxy-α-ionone in vitro. MnCCD1 could also cleave lycopene and β-carotene at the 9, 10 (9', 10') bind bond to produce pseudoionone and β-ionone, respectively, in E. coli accumulating carotenoids. The enzyme activity of MnCCD1 was reached 2.98 U/mL at optimized conditions (temperature 28 °C, IPTG 0.1 mM, induction time 24 h). The biochemical characterization of MnCCD1 revealed the optimal activities were at pH 8.4 and 35 °C. The addition of 10 % ethanol could increase enzyme activity at above 15 %. However, an obvious decline was observed on enzyme activity as the concentration of Fe 2 + increased (0-1 mM). The V max for β-apo-8'-carotenal was 72.5 U/mg, while the K m was 0.83 mM. The results provide a foundation for developing the application of carotenoid cleavage dioxygenases as biocatalysis and synthetic biology platforms to produce volatile aroma components from carotenoids.

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

Cited by 32 publications

References 43 publications

β-Ionone: Its Occurrence and Biological Function and Metabolic Engineering

β-Ionone: Its Occurrence and Biological Function and Metabolic Engineering

Characterization of a Novel Lutein Cleavage Dioxygenase, EhLCD, from Enterobacter hormaechei YT-3 for the Enzymatic Synthesis of 3-Hydroxy-β-ionone from Lutein

Overexpression and Characterization of a Novel Plant Carotenoid Cleavage Dioxygenase 1 from Morus notabilis

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