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Metadados | Descrição | Idioma |
---|---|---|
Autor(es): dc.contributor | Universidade Estadual de Campinas (UNICAMP) | - |
Autor(es): dc.contributor | Universidade Estadual Paulista (Unesp) | - |
Autor(es): dc.contributor | Universidade de São Paulo (USP) | - |
Autor(es): dc.creator | Rubio, Marcelo Ventura | - |
Autor(es): dc.creator | Terrasan, César Rafael Fanchini | - |
Autor(es): dc.creator | Contesini, Fabiano Jares | - |
Autor(es): dc.creator | Zubieta, Mariane Paludetti | - |
Autor(es): dc.creator | Gerhardt, Jaqueline Aline | - |
Autor(es): dc.creator | Oliveira, Leandro Cristante [UNESP] | - |
Autor(es): dc.creator | De Souza Schmidt Gonçalves, Any Elisa | - |
Autor(es): dc.creator | Almeida, Fausto | - |
Autor(es): dc.creator | Smith, Bradley Joseph | - |
Autor(es): dc.creator | De Souza, Gustavo Henrique Martins Ferreira | - |
Autor(es): dc.creator | Dias, Artur Hermano Sampaio | - |
Autor(es): dc.creator | Skaf, Munir | - |
Autor(es): dc.creator | Damasio, André | - |
Data de aceite: dc.date.accessioned | 2022-02-22T00:32:33Z | - |
Data de disponibilização: dc.date.available | 2022-02-22T00:32:33Z | - |
Data de envio: dc.date.issued | 2020-12-11 | - |
Data de envio: dc.date.issued | 2020-12-11 | - |
Data de envio: dc.date.issued | 2019-11-13 | - |
Fonte completa do material: dc.identifier | http://dx.doi.org/10.1186/s13068-019-1609-2 | - |
Fonte completa do material: dc.identifier | http://hdl.handle.net/11449/201085 | - |
Fonte: dc.identifier.uri | http://educapes.capes.gov.br/handle/11449/201085 | - |
Descrição: dc.description | Background: β-Xylosidases are glycoside hydrolases (GHs) that cleave xylooligosaccharides and/or xylobiose into shorter oligosaccharides and xylose. Aspergillus nidulans is an established genetic model and good source of carbohydrate-active enzymes (CAZymes). Most fungal enzymes are N-glycosylated, which influences their secretion, stability, activity, signalization, and protease protection. A greater understanding of the N-glycosylation process would contribute to better address the current bottlenecks in obtaining high secretion yields of fungal proteins for industrial applications. Results: In this study, BxlB-a highly secreted GH3 β-xylosidase from A. nidulans, presenting high activity and several N-glycosylation sites-was selected for N-glycosylation engineering. Several glycomutants were designed to investigate the influence of N-glycans on BxlB secretion and function. The non-glycosylated mutant (BxlBnon-glyc) showed similar levels of enzyme secretion and activity compared to the wild-type (BxlBwt), while a partially glycosylated mutant (BxlBN1;5;7) exhibited increased activity. Additionally, there was no enzyme secretion in the mutant in which the N-glycosylation context was changed by the introduction of four new N-glycosylation sites (BxlBCC), despite the high transcript levels. BxlBwt, BxlBnon-glyc, and BxlBN1;5;7 formed similar secondary structures, though the mutants had lower melting temperatures compared to the wild type. Six additional glycomutants were designed based on BxlBN1;5;7, to better understand its increased activity. Among them, the two glycomutants which maintained only two N-glycosylation sites each (BxlBN1;5 and BxlBN5;7) showed improved catalytic efficiency, whereas the other four mutants' catalytic efficiencies were reduced. The N-glycosylation site N5 is important for improved BxlB catalytic efficiency, but needs to be complemented by N1 and/or N7. Molecular dynamics simulations of BxlBnon-glyc and BxlBN1;5 reveals that the mobility pattern of structural elements in the vicinity of the catalytic pocket changes upon N1 and N5 N-glycosylation sites, enhancing substrate binding properties which may underlie the observed differences in catalytic efficiency between BxlBnon-glyc and BxlBN1;5. Conclusions: This study demonstrates the influence of N-glycosylation on A. nidulans BxlB production and function, reinforcing that protein glycoengineering is a promising tool for enhancing thermal stability, secretion, and enzymatic activity. Our report may also support biotechnological applications for N-glycosylation modification of other CAZymes. | - |
Descrição: dc.description | Department of Biochemistry and Tissue Biology Institute of Biology University of Campinas (UNICAMP), Rua Monteiro Lobato, 255 Cidade Universitária Zeferino Vaz | - |
Descrição: dc.description | Department of Physics Institute of Biosciences Humanities and Exact Sciences São Paulo State University (UNESP) | - |
Descrição: dc.description | Department of Medical Science Faculty of Medicine University of Campinas (UNICAMP) | - |
Descrição: dc.description | Department of Biochemistry and Immunology Ribeirão Preto Medical School University of São Paulo (USP) | - |
Descrição: dc.description | Institute of Chemistry and Center for Computing in Engineering and Sciences University of Campinas (UNICAMP) | - |
Descrição: dc.description | Department of Physics Institute of Biosciences Humanities and Exact Sciences São Paulo State University (UNESP) | - |
Idioma: dc.language | en | - |
Relação: dc.relation | Biotechnology for Biofuels | - |
???dc.source???: dc.source | Scopus | - |
Palavras-chave: dc.subject | Aspergillus nidulans | - |
Palavras-chave: dc.subject | CAZyme | - |
Palavras-chave: dc.subject | Enzyme secretion | - |
Palavras-chave: dc.subject | Glycomutants | - |
Palavras-chave: dc.subject | Glycoside hydrolase family 3 | - |
Palavras-chave: dc.subject | N-glycosylation | - |
Palavras-chave: dc.subject | β-Xylosidase | - |
Título: dc.title | Redesigning N-glycosylation sites in a GH3 β-xylosidase improves the enzymatic efficiency | - |
Tipo de arquivo: dc.type | livro digital | - |
Aparece nas coleções: | Repositório Institucional - Unesp |
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