Enhancing thermostability by modifying flexible surface loops in an evolved high-redox potential laccase
| Autor: | Fátima Lucas, Ana I. Vicente, Javier Viña-Gonzalez, Victor Guallar, Miguel Alcalde, Emanuele Monza, Ivan Mateljak |
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| Přispěvatelé: | European Commission, Ministerio de Economía y Competitividad (España), Alcalde Galeote, Miguel [0000-0001-6780-7616], Alcalde Galeote, Miguel |
| Rok vydání: | 2019 |
| Předmět: |
Laccase
Environmental Engineering Chemistry General Chemical Engineering B-factor 02 engineering and technology Molecular dynamics 021001 nanoscience & nanotechnology Flexible loops Redox MORPHING High-redox potential laccase 020401 chemical engineering Chemical engineering B factor 0204 chemical engineering Thermostability 0210 nano-technology Biotechnology |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname |
| Popis: | [EN] High-redox potential laccases (HRPLs) from white-rot fungi are versatile biocatalysts whose practical use is highly dependent on their thermostability. In this work, an evolved HRPL variant was subjected to structure-guided evolution to improve its thermostability. We first selected several surface flexible loops in the laccase structure by inspecting them through molecular dynamics and an analysis of B-factors. The resulting segments were grouped into three MORPHING (Mutagenic Organized Recombination Process by Homologous In vivo Grouping) blocks, which were constructed in Saccharomyces cerevisiae and explored at high temperatures. This evolution process gave rise to a double mutant that showed a half-life at 70°C enhanced by 31 min with an optimum temperature for activity of 75°C and similar kinetic parameters. The Ser264Lys and Ser356Asn mutations modified the contacts established between these residues and those that surround them, altering the surface loops and thereby the enzyme properties. FP7 Food, Agriculture and Fisheries, Biotechnology, Grant/Award Numbers: FP7‐KBBE‐2013‐7‐613549‐INDOX, COST‐ActionCM1303; Ministerio de Ciencia e Innovación, Grant/Award Numbers: BIO2016‐79106‐R‐Lignolution, CTQ2016‐79138‐R |
| Databáze: | OpenAIRE |
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