Biotransformation of Isoflavone Using Enzymatic Reactions
Abstract
:1. Introduction
2. Results and Discussion
2.1. Chemical Structure of 7,3',4'-Trihydroxyisoflavone from Daidzein
2.2. Identification and Functional Construction of CYP105D7 and Redox Partner in E. coli System
2.3. Overexpression and Purification of CYP105D7, PdR and Pdx
2.4. Characterization of Recombinant CYP105D7 and Kinetic Parameter
Substrate | Km [μM] | kcat [min−1] | kcat/Km [μM−1/min−1] * |
---|---|---|---|
Daidzein | 21.83 ± 6.3 | 15.01 ± 0.6 | 0.69 |
2.5. Hydroxylation of Daidzein by CYP105D7 in Vivo and in Vitro System
2.6. Effect of P450 Inhibitors on the Hydroxylation of Daidzein
P450 inhibitor * | Reactive activity (%) |
---|---|
None | 100 |
Coumarin | 2 |
Erythromycin | 63 |
Ketoconazole | 14 |
3. Experimental
3.1. Chemicals
3.2. Plasmid Construction
3.3. Cloning of CYP105D7 from Streptomyces avermitilis MA4680
3.4. Coexpression of CYP105D7 and Redox Partner for in Vivo System
3.5. Purification of CYP105D7 and Redox Partner for in Vitro System
3.6. CYP105D7 Concentration Measurement and Absorption Spectra
3.7. Kinetic Study for Substrate Activity with CYP105D7
3.8. Daidzein Hydroxylation in Vivo and in Vitro System
3.9. High Performance Liquid Chromatography (HPLC) Analysis of Product
3.10. Gas Chromatography (GC)/Mass Spectrometry (MS) Analysis
3.11. Effects of P450 Inhibitors on Hydroxylation of Daidzein
4. Conclusions
Acknowledgments
References
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Roh, C. Biotransformation of Isoflavone Using Enzymatic Reactions. Molecules 2013, 18, 3028-3040. https://doi.org/10.3390/molecules18033028
Roh C. Biotransformation of Isoflavone Using Enzymatic Reactions. Molecules. 2013; 18(3):3028-3040. https://doi.org/10.3390/molecules18033028
Chicago/Turabian StyleRoh, Changhyun. 2013. "Biotransformation of Isoflavone Using Enzymatic Reactions" Molecules 18, no. 3: 3028-3040. https://doi.org/10.3390/molecules18033028