Functional Expression and One-Step Protein Purification of Manganese Peroxidase 1 (rMnP1) from Phanerochaete chrysosporium Using the E. coli -Expression System.
Autor: | Pech-Canul AC; CONACyT-Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., Carrillo-Campos J; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., Ballinas-Casarrubias ML; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., Solis-Oviedo RL; Healthy Life Importadora, Exportadora Y Productora De Alimentos Y Bebidas, Sociedad De Responsabilidad Limitada De Capital Variable, Chihuahua 31125, Mexico., Hernández-Rascón SK; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., Hernández-Ochoa LR; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., Gutiérrez-Méndez N; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico., García-Triana A; Faculty of Chemical Sciences, Autonomous University of Chihuahua, Campus II, Chihuahua 31125, Mexico. |
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Jazyk: | angličtina |
Zdroj: | International journal of molecular sciences [Int J Mol Sci] 2020 Jan 09; Vol. 21 (2). Date of Electronic Publication: 2020 Jan 09. |
DOI: | 10.3390/ijms21020416 |
Abstrakt: | Manganese peroxidases (MnP) from the white-rot fungi Phanerochaete chrysosporium catalyse the oxidation of Mn 2+ to Mn 3+ , a strong oxidizer able to oxidize a wide variety of organic compounds. Different approaches have been used to unravel the enzymatic properties and potential applications of MnP. However, these efforts have been hampered by the limited production of native MnP by fungi. Heterologous expression of MnP has been achieved in both eukaryotic and prokaryotic expression systems, although with limited production and many disadvantages in the process. Here we described a novel molecular approach for the expression and purification of manganese peroxidase isoform 1 (MnP1) from P. chrysosporium using an E. coli -expression system. The proposed strategy involved the codon optimization and chemical synthesis of the MnP1 gene for optimised expression in the E. coli T7 shuffle host. Recombinant MnP1 (rMnP1) was expressed as a fusion protein, which was recovered from solubilised inclusion bodies. rMnP1 was purified from the fusion protein using intein-based protein purification techniques and a one-step affinity chromatography. The designated strategy allowed production of an active enzyme able to oxidize guaiacol or Mn 2+ . |
Databáze: | MEDLINE |
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