Use este identificador para citar ou linkar para este item: https://locus.ufv.br//handle/123456789/21788
Tipo: Artigo
Título: Thermostability improvement of Orpinomyces sp. xylanase by directed evolution
Autor(es): Trevizano, Larissa Mattos
Ventorim, Rafaela Zandonade
Rezende, Sebastião Tavares de
Silva Junior, Floriano Paes
Guimarães, Valéria Monteze
Abstract: The methodology of directed evolution, using the mutagenic technique of error-prone PCR has been used to improve the thermostability of enzymes. This method was applied to the endo-β-1,4-xylanase from Orpinomyces strain PC-2. The constructed library of xylanase (xynA) mutants was subjected to several screening cycles in plates with azo-xylan-agarose as substrate and four thermostable mutants (M1–M4) were selected. Homology models for these thermostable mutants were constructed to identify the location of the residues changed by error-prone PCR and to investigate the effect of these mutations on the xylanase properties. Xylanase activities of the mutants and wild type were maximal at 60 °C and in the pH range of 5–7. The mutants displayed higher thermostability than the wild type XynA, where the wild type showed a half-life at 60 °C of 7.92 min, while half-life values for M1, M2, M3 and M4 were 209, 33.2, 401 and 15.3 min, respectively. Additionally, M3 and M4 presented a good performance in more extreme pH conditions. The mutants retained their ability to hydrolyze birchwood and oat spelt xylans, which are substrates presenting different degrees of branching.
Palavras-chave: Error-prone PCR
Xylanase
Thermostability
Orpinomyces
Editor: Journal of Molecular Catalysis B: Enzymatic
Tipo de Acesso: Elsevier B.V.
URI: https://doi.org/10.1016/j.molcatb.2012.04.021
http://www.locus.ufv.br/handle/123456789/21788
Data do documento: Set-2012
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