Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation
Bacterial cytochrome P450 (CYP) enzymes are versatile biocatalysts that are responsible for the biotransformation of diverse endogenous substances. CYP105A5 from <i>Streptomyces</i> sp. showed substrate flexibility with different flavonoids and was able to catalyze O-demethylation of bio...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-10-01
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| Series: | Catalysts |
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| Online Access: | https://www.mdpi.com/2073-4344/12/10/1157 |
| _version_ | 1797474460324855808 |
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| author | Pradeep Subedi Jong Kook Park Tae-Jin Oh |
| author_facet | Pradeep Subedi Jong Kook Park Tae-Jin Oh |
| author_sort | Pradeep Subedi |
| collection | DOAJ |
| description | Bacterial cytochrome P450 (CYP) enzymes are versatile biocatalysts that are responsible for the biotransformation of diverse endogenous substances. CYP105A5 from <i>Streptomyces</i> sp. showed substrate flexibility with different flavonoids and was able to catalyze O-demethylation of biochanin A, regioselective C3′-hydroxylation of daidzein, genistein, and naringenin, and additional C8-hydroxylation for daidzein using heterologous redox partners putidaredoxin and putidaredoxin reductase. By rational design of substrate-binding pocket based on experimental data, homology modeling, and molecular docking analysis, we enhanced the product formation rate of flavonoids. The double mutant L100A/I302A and L100A/I408N exhibited greatly enhanced in vivo conversion rates for flavonoid hydroxylation. Particularly, the L100A/I302A mutant’s <i>k<sub>cat</sub>/K<sub>m</sub></i> values and in vivo conversion rate increased by 1.68-fold and 2.57-fold, respectively, for naringenin. Overall, our result might facilitate the potential use of CYP105A5 for future modification and application in whole-cell biocatalysts for the production of valuable polyphenols. |
| first_indexed | 2024-03-09T20:30:25Z |
| format | Article |
| id | doaj.art-b8ecdc7e345c4bbab188f472e21035ca |
| institution | Directory Open Access Journal |
| issn | 2073-4344 |
| language | English |
| last_indexed | 2024-03-09T20:30:25Z |
| publishDate | 2022-10-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Catalysts |
| spelling | doaj.art-b8ecdc7e345c4bbab188f472e21035ca2023-11-23T23:24:36ZengMDPI AGCatalysts2073-43442022-10-011210115710.3390/catal12101157Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid HydroxylationPradeep Subedi0Jong Kook Park1Tae-Jin Oh2Department of Life Science and Biochemical Engineering, Sunmoon University, 70 Sunmoon-ro 221, Tangjeong-myeon, Asan-si 31460, KoreaDepartment of Biomedical Science and Research Institute for Bioscience & Biotechnology, Hallym University, 1 Hallymdaehak-gil, Chuncheon 24252, KoreaDepartment of Life Science and Biochemical Engineering, Sunmoon University, 70 Sunmoon-ro 221, Tangjeong-myeon, Asan-si 31460, KoreaBacterial cytochrome P450 (CYP) enzymes are versatile biocatalysts that are responsible for the biotransformation of diverse endogenous substances. CYP105A5 from <i>Streptomyces</i> sp. showed substrate flexibility with different flavonoids and was able to catalyze O-demethylation of biochanin A, regioselective C3′-hydroxylation of daidzein, genistein, and naringenin, and additional C8-hydroxylation for daidzein using heterologous redox partners putidaredoxin and putidaredoxin reductase. By rational design of substrate-binding pocket based on experimental data, homology modeling, and molecular docking analysis, we enhanced the product formation rate of flavonoids. The double mutant L100A/I302A and L100A/I408N exhibited greatly enhanced in vivo conversion rates for flavonoid hydroxylation. Particularly, the L100A/I302A mutant’s <i>k<sub>cat</sub>/K<sub>m</sub></i> values and in vivo conversion rate increased by 1.68-fold and 2.57-fold, respectively, for naringenin. Overall, our result might facilitate the potential use of CYP105A5 for future modification and application in whole-cell biocatalysts for the production of valuable polyphenols.https://www.mdpi.com/2073-4344/12/10/1157biocatalysiscytochrome P450 monooxygenaseflavonoid O-demethylation/hydroxylationprotein engineeringwhole-cell biotransformation |
| spellingShingle | Pradeep Subedi Jong Kook Park Tae-Jin Oh Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation Catalysts biocatalysis cytochrome P450 monooxygenase flavonoid O-demethylation/hydroxylation protein engineering whole-cell biotransformation |
| title | Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation |
| title_full | Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation |
| title_fullStr | Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation |
| title_full_unstemmed | Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation |
| title_short | Engineering of Microbial Substrate Promiscuous CYP105A5 for Improving the Flavonoid Hydroxylation |
| title_sort | engineering of microbial substrate promiscuous cyp105a5 for improving the flavonoid hydroxylation |
| topic | biocatalysis cytochrome P450 monooxygenase flavonoid O-demethylation/hydroxylation protein engineering whole-cell biotransformation |
| url | https://www.mdpi.com/2073-4344/12/10/1157 |
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