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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 |
Aparece nas coleções: | Artigos |
Arquivos associados a este item:
Arquivo | Descrição | Tamanho | Formato | |
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artigo.pdf Until 2100-12-31 | texto completo | 592,76 kB | Adobe PDF | Visualizar/Abrir ACESSO RESTRITO |
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