Engineering of a fungal laccase to develop a robust, versatile and highly-expressed biocatalyst for sustainable chemistry
| Autor: | Jesper Vind, Gerard Santiago, Sibel Kilic, Joan Gilabert, Ángel T. Martínez, Victor Guallar, Felipe de Salas, Susana Camarero, Pablo Aza, Mehmet E. Sener |
|---|---|
| Přispěvatelé: | European Commission, Ministerio de Economía y Competitividad (España), Ministerio de Ciencia, Innovación y Universidades (España), Consejo Superior de Investigaciones Científicas (España), De Salas, Felipe, Gilabert, Joan F., Santiago, Gerard, Vind, Jesper, Guallar, Victor, Martínez, Angel T., Camarero, Susana, De Salas, Felipe [0000-0002-0057-0180], Gilabert, Joan F. [0000-0002-5008-5482], Santiago, Gerard [0000-0002-0506-3049], Vind, Jesper [0000-0002-1336-9531], Guallar, Victor [0000-0002-4580-1114], Martínez, Angel T. [0000-0002-1584-2863], Camarero, Susana [0000-0002-2812-895X] |
| Rok vydání: | 2019 |
| Předmět: |
Green chemistry
Molecular-cloning 010402 general chemistry 01 natural sciences C-terminus chemistry.chemical_compound Aniline Aspergillus oryzae Polyaniline Environmental Chemistry Enzyme kinetics Laccase biology 010405 organic chemistry Substrate (chemistry) Directed evolution biology.organism_classification Pollution Combinatorial chemistry Melanocarpus-albomyces 0104 chemical sciences Crystal-structure chemistry Mutagenesis Stability Catalytic-properties |
| Zdroj: | Green Chemistry Digital.CSIC. Repositorio Institucional del CSIC instname |
| ISSN: | 1463-9270 1463-9262 |
| DOI: | 10.1039/c9gc02475a |
| Popis: | 12 p.-12 fig.-1 tab. Fungal laccases can play an important role as biocatalysts in organic chemistry to replace chemical synthesis. In a previous work we synthesized conductive polyaniline using a high-redox potential laccase from our collection of recombinant fungal variants. Still, the oxidation of aniline is hindered by the reaction conditions (low pH and presence of anionic surfactants). Thus, we tackle here the directed evolution of the enzyme assisted by computational simulation aiming at improving aniline oxidation at the required polymerization conditions while maintaining the enzyme's substrate promiscuity. Simultaneously, its secretion by the host used for the engineering (Saccharomyces cerevisiae) was enhanced. Then, the improved laccase variant was overproduced in the industrial host Aspergillus oryzae and assayed for one-pot synthesis of polyaniline and naphtol-derived dyes whose textile dyeing properties were verified in an industrial environment. Finally, modification of its C-terminal tail further enhanced laccase stability by flexibilization of the region. The resulting biocatalyst displays noticeable stability at high temperature and extreme pH while shows improved k(cat) values on the different substrates tested. Moreover, it is remarkably produced in S. cerevisiae at rates not formerly reported in the literature. These facts, together with the overexpression in A. oryzae opens new scenarios for its further development and application. This work has been funded by the INDOX EU project (KBBE- 2013-7-613549), the Spanish projects BIO2017-86559-R and CTQ2016-79138-R and the BBI JU project WoodZymes (H2020-BBI-JU-792070). We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI). |
| Databáze: | OpenAIRE |
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