Colorimetric High-Throughput Screening Assays for the Directed Evolution of Fungal Laccases
In this chapter we describe several high-throughput screening assays for the evaluation of mutant libraries for the directed evolution of fungal laccases in the yeast Saccharomyces cerevisiae. The assays are based on the direct oxidation of three syringyl-type phenols derived from lignin (sinapic ac...
| Autores: | , |
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| Tipo de recurso: | otro |
| Estado: | Versión publicada |
| Fecha de publicación: | 2017 |
| País: | España |
| Institución: | Consejo Superior de Investigaciones Científicas (CSIC) |
| Repositorio: | DIGITAL.CSIC. Repositorio Institucional del CSIC |
| OAI Identifier: | oai:dnet:digitalcsic_::f8157c2cd5ed5d4916dcec66566a29e2 |
| Acceso en línea: | http://hdl.handle.net/10261/427664 https://api.elsevier.com/content/abstract/scopus_id/85032641183 |
| Access Level: | acceso abierto |
| Palabra clave: | Colorimetric assays Directed evolution High-throughput screening Laccase Mutant libraries Synthetic organic dyes Syringyl-type phenols Violuric acid |
| Sumario: | In this chapter we describe several high-throughput screening assays for the evaluation of mutant libraries for the directed evolution of fungal laccases in the yeast Saccharomyces cerevisiae. The assays are based on the direct oxidation of three syringyl-type phenols derived from lignin (sinapic acid, acetosyringone, and syringaldehyde), an artificial laccase mediator (violuric acid), and three organic synthetic dyes (Methyl Orange, Evans Blue, and Remazol Brilliant Blue). While the assays with the natural phenols can be used for laccases with low redox potential, the rest are exclusive for high-redox potential laccases. In fact, the violuric acid assay is devised as a method to ascertain that the high-redox potential of laccase is not lost during directed evolution. |
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