Characterization of a Novel Lutein Cleavage Dioxygenase, EhLCD, from <i>Enterobacter hormaechei</i> YT-3 for the Enzymatic Synthesis of 3-Hydroxy-<i>β</i>-ionone from Lutein
3-Hydroxy-<i>β</i>-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...
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| Autores principales: | , , , , , , , , |
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| Formato: | article |
| Lenguaje: | EN |
| Publicado: |
MDPI AG
2021
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| Materias: | |
| Acceso en línea: | https://doaj.org/article/d8edddf174ce4afab1258f0f07765ff7 |
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| Sumario: | 3-Hydroxy-<i>β</i>-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 <i>Enterobacter hormaechei</i> YT-3 to convert lutein to 3-hydroxy-<i>β</i>-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, <i>β</i>-cryptoxanthin, and <i>β</i>-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 O<sub>2</sub> into 3-hydroxy-<i>β</i>-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 Fe<sup>2+</sup>-dependent protein with 1 molar equivalent of non-haem Fe<sup>2+</sup>. 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-<i>β</i>-ionone production were 60 μM lutein/L, 1.5 U/mL, and 2% (<i>w</i>/<i>v</i>), respectively. Under optimum conditions, EhLCD produced 3-hydroxy-<i>β</i>-ionone (637.2 mg/L) in 60 min with a conversion of 87.0% (<i>w</i>/<i>w</i>), indicating that this enzyme is a potential candidate for the enzymatic synthesis of 3-hydroxy-<i>β</i>-ionone in biotechnological applications. |
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