Identification of a self-sufficient cytochrome P450 monooxygenase from Cupriavidus pinatubonensis JMP134 involved in 2-hydroxyphenylacetic acid catabolism, via homogentisate pathway
| Autor: | Daniela Ruiz, José Eduardo González-Pastor, Bernardo González, Pamela Villegas, Carla Gárate-Castro, Danilo Pérez-Pantoja, Víctor de Lorenzo, Raúl Donoso |
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| Přispěvatelé: | De Lorenzo, V. [0000-0002-6041-2731], Pérez Pantoja, D. [0000-0001-5720-2162], Comunidad de Madrid, Fondo Nacional de Desarrollo Científico y Tecnológico (FONDECYT) |
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Oxygenase
Operon Bioengineering Cytochrome P450 Applied Microbiology and Biotechnology Biochemistry 03 medical and health sciences Cytochrome P-450 Enzyme System Gene 030304 developmental biology Homogentisate 1 2-dioxygenase Phenylacetates 0303 health sciences biology 030306 microbiology Catabolism Chemistry Cupriavidus pinatubonensis JMP134 Cupriavidus Monooxygenase biology.protein Heterologous expression TP248.13-248.65 Biotechnology |
| Zdroj: | Microbial Biotechnology, Vol 14, Iss 5, Pp 1944-1960 (2021) DIGITAL.INTA Repositorio Digital del Instituto Nacional de Técnica Aeroespacial instname |
| DOI: | 10.1111/1751-7915.13865 |
| Popis: | The self-sufficient cytochrome P450 RhF and its homologues belonging to the CYP116B subfamily have attracted considerable attention due to the potential for biotechnological applications based in their ability to catalyse an array of challenging oxidative reactions without requiring additional protein partners. In this work, we showed for the first time that a CYP116B self-sufficient cytochrome P450 encoded by the ohpA gene harboured by Cupriavidus pinatubonensis JMP134, a β-proteobacterium model for biodegradative pathways, catalyses the conversion of 2-hydroxyphenylacetic acid (2-HPA) into homogentisate. Mutational analysis and HPLC metabolite detection in strain JMP134 showed that 2-HPA is degraded through the well-known homogentisate pathway requiring a 2-HPA 5-hydroxylase activity provided by OhpA, which was additionally supported by heterologous expression and enzyme assays. The ohpA gene belongs to an operon including also ohpT, coding for a substrate-binding subunit of a putative transporter, whose expression is driven by an inducible promoter responsive to 2-HPA in presence of a predicted OhpR transcriptional regulator. OhpA homologues can be found in several genera belonging to Actinobacteria and α-, β- and γ-proteobacteria lineages indicating a widespread distribution of 2-HPA catabolism via homogentisate route. These results provide first time evidence for the natural function of members of the CYP116B self-sufficient oxygenases and represent a significant input to support novel kinetic and structural studies to develop cytochrome P450-based biocatalytic processes. This work was funded by FONDECYT 1201741, ANID PIA/Anillo ACT172128, and ANID PIA/BASAL FB0002 grants, the InGEMICS-CM (S2017/BMD-3691) Project of the Comunidad de Madrid – European Structural and Investment Funds – (FSE, FECER), and the L318-07 Project supported by the Fund of Scientific and Technological Equipment, year 2018, Universidad Tecnológica Metropolitana. Peerreview |
| Databáze: | OpenAIRE |
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