Computational-Aided Engineering of a Selective Unspecific Peroxygenase toward Enantiodivergent β-Ionone Hydroxylation
Unspecific peroxygenases (UPOs) perform oxyfunctionalizations for a wide range of substrates utilizing H2O2 without the need for further reductive equivalents or electron transfer chains. Tailoring these promising enzymes toward industrial application was intensely pursued in the last decade with en...
| Autores: | , , , , , |
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| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2023 |
| País: | España |
| Institución: | Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya) |
| Repositorio: | Recercat. Dipósit de la Recerca de Catalunya |
| OAI Identifier: | oai:recercat.cat:10256/23576 |
| Acceso en línea: | http://hdl.handle.net/10256/23576 |
| Access Level: | acceso abierto |
| Palabra clave: | Pèptids Peptides Proteïnes Proteins Reaccions químiques regioselectives Chemical reactions |
| Sumario: | Unspecific peroxygenases (UPOs) perform oxyfunctionalizations for a wide range of substrates utilizing H2O2 without the need for further reductive equivalents or electron transfer chains. Tailoring these promising enzymes toward industrial application was intensely pursued in the last decade with engineering campaigns addressing the heterologous expression, activity, stability, and improvements in chemo- and regioselectivity. One hitherto missing integral part was the targeted engineering of enantioselectivity for specific substrates with poor starting enantioselectivity. In this work, we present the engineering of the short-type MthUPO toward the enantiodivergent hydroxylation of the terpene model substrate, β-ionone. Guided by computational modeling, we designed a small smart library and screened it with a GC–MS setup. After two rounds of iterative protein evolution, the activity increased up to 17-fold and reached a regioselectivity of up to 99.6% for the 4-hydroxy-β-ionone. Enantiodivergent variants were identified with enantiomeric ratios of 96.6:3.4 (R) and 0.3:99.7 (S), respectively |
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