Cloning and heterologous expression of P450Lent4B11, a novel bacterial P450 gene, for hydroxylation of an antifungal agent sordaricin
| Autor: | Motoo Kobayashi, Akihiko Fujie, Motoi Ueno, Takashi Shibata |
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| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Antifungal Agents Stereochemistry 030106 microbiology Hydroxylation 01 natural sciences 03 medical and health sciences chemistry.chemical_compound Cytochrome P-450 Enzyme System Drug Discovery Actinomycetales Moiety Cloning Molecular Biotransformation Pharmacology Fungal protein Natural product biology 010405 organic chemistry Fungi Cytochrome P450 Metyrapone 0104 chemical sciences Elongation factor Aglycone chemistry Indenes Genes Bacterial biology.protein Heterologous expression Diterpenes Oxidation-Reduction |
| Zdroj: | The Journal of antibiotics. 73(9) |
| ISSN: | 1881-1469 |
| Popis: | Microbial transformation is known to be one of promising options to add functional groups such as a hydroxyl moiety to active base compounds to generate their derivatives. Sordaricin, a diterpene aglycone of the natural product sordarin, is an antifungal agent to selectively inhibit fungal protein synthesis by stabilizing the ribosome/EF-2 (elongation factor 2) complex. We screened actinomycetes to catalyze hydroxylation of sordaricin on the basis that the hydroxyl moiety would make it easier to generate derivatives of sordaricin. As a result of the screening, 6-hydroxylation of sordaricin was found to be catalyzed by Lentzea sp. 7887. We found that the cytochrome P450 inhibitor metyrapone inhibited this reaction, suggesting that a cytochrome P450 may be responsible for the biotransformation. As a next step, we cloned multiple cytochrome P450 genes, one of which were named P450Lent4B11, using degenerate PCR primers. The expressed cytochrome P450 derived from the P450Lent4B11 gene provided a different absorbance spectrum pattern from original one when it was incubated with sordaricin. Moreover, in cell-free conditions, the corresponding cytochrome P450 displayed the 6-hydroxylation activity toward sordaricin. Taken together, these results indicate that P450Lent4B11, derived from Lentzea sp. 7887, should be responsible for catalyzing 6-hydroxylation of sordaricin. |
| Databáze: | OpenAIRE |
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