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Title

Laboratory Evolution of High-Redox Potential Laccases

AuthorsMaté, Diana M. ; García-Burgos, Carlos ; García-Ruiz, Eva ; Ballesteros Olmo, Antonio ; Camarero, Susana ; Alcalde Galeote, Miguel
KeywordsLacasse
Chemistry
Directed Molecular Evolutions
Enzymology
Genetics
Metabolism
Oxidation Reduction Reaction
pH
Protein Stability
Protein Tertiary Structure
Saccharomyces Cerevisae
Site Directed Mutagenesis
Issue Date24-Sep-2010
PublisherElsevier
CitationChemistry and Biology 17(9) : 1030-1041 (2010)
AbstractThermostable laccases with a high-redox potential have been engineered through a strategy that combines directed evolution with rational approaches. The original laccase signal sequence was replaced by the α-factor prepro-leader, and the corresponding fusion gene was targeted for joint laboratory evolution with the aim of improving kinetics and secretion by Saccharomyces cerevisiae, while retaining high thermostability. After eight rounds of molecular evolution, the total laccase activity was enhanced 34,000-fold culminating in the OB-1 mutant as the last variant of the evolution process, a highly active and stable enzyme in terms of temperature, pH range, and organic cosolvents. Mutations in the hydrophobic core of the evolved α-factor prepro-leader enhanced functional expression, whereas some mutations in the mature protein improved its catalytic capacities by altering the interactions with the surrounding residues
Description12 páginas, 5 figuras, 2 tablas -- PAGS nros. 1030-1041
Publisher version (URL)http://dx.doi.org/10.1016/j.chembiol.2010.07.010
URIhttp://hdl.handle.net/10261/46385
DOI10.1016/j.chembiol.2010.07.010
ISSN1074-5521
E-ISSN1879-1301
Appears in Collections:(CIB) Artículos
(ICP) Artículos
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