Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii
Autor(a) principal: | |
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Data de Publicação: | 2010 |
Outros Autores: | , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/1822/11325 |
Resumo: | To explore the potential of Ashbya gossypii as a host for the expression of recombinant proteins and to assess whether protein secretion would be more similar to the closely related Saccharomyces cerevisiae or to other filamentous fungi, endoglucanase I (EGI) and cellobiohydrolase I (CBHI) from the fungus Trichoderma reesei were successfully expressed in A. gossypii from plasmids containing the two micron sequences from S. cerevisiae, under the S. cerevisiae PGK1 promoter. The native signal sequences of EGI and CBHI were able to direct the secretion of EGI and CBHI into the culture medium in A. gossypii. Although CBHI activity was not detected using 4- methylumbelliferyl-β-D-lactoside as substrate, the protein was detected by Western blot using monoclonal antibodies. EGI activity was detectable, the specific activity being comparable to that produced by a similar EGI producing S. cerevisiae construct. More EGI was secreted than CBHI, or more active protein was produced. Partial characterization of CBHI and EGI expressed in A. gossypii revealed overglycosylation when compared with the native T. reesei proteins, but the glycosylation was less extensive than on cellulases expressed in S. cerevisiae. |
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Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypiiAshbya gossypiiRecombinant protein productionTrichoderma reesei endoglucanase ITrichoderma reesei cellobiohydrolase ICellulases heterologous expressionScience & TechnologyTo explore the potential of Ashbya gossypii as a host for the expression of recombinant proteins and to assess whether protein secretion would be more similar to the closely related Saccharomyces cerevisiae or to other filamentous fungi, endoglucanase I (EGI) and cellobiohydrolase I (CBHI) from the fungus Trichoderma reesei were successfully expressed in A. gossypii from plasmids containing the two micron sequences from S. cerevisiae, under the S. cerevisiae PGK1 promoter. The native signal sequences of EGI and CBHI were able to direct the secretion of EGI and CBHI into the culture medium in A. gossypii. Although CBHI activity was not detected using 4- methylumbelliferyl-β-D-lactoside as substrate, the protein was detected by Western blot using monoclonal antibodies. EGI activity was detectable, the specific activity being comparable to that produced by a similar EGI producing S. cerevisiae construct. More EGI was secreted than CBHI, or more active protein was produced. Partial characterization of CBHI and EGI expressed in A. gossypii revealed overglycosylation when compared with the native T. reesei proteins, but the glycosylation was less extensive than on cellulases expressed in S. cerevisiae.Fundação para a Ciência e a Tecnologia (FCT)Springer VerlagUniversidade do MinhoRibeiro, OrquídeaWiebe, MarilynIlmén, MarjaDomingues, LucíliaPenttilä, Merja20102010-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/1822/11325eng"Applied Microbiology and Biotechnology". ISSN 0175-7598. 87:4 (2010) 1437-1446.0175-759810.1007/s00253-010-2610-720422178info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-07-21T12:15:02ZPortal AgregadorONG |
dc.title.none.fl_str_mv |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
title |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
spellingShingle |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii Ribeiro, Orquídea Ashbya gossypii Recombinant protein production Trichoderma reesei endoglucanase I Trichoderma reesei cellobiohydrolase I Cellulases heterologous expression Science & Technology |
title_short |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
title_full |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
title_fullStr |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
title_full_unstemmed |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
title_sort |
Expression of Trichoderma reesei cellulases CBHI and EGI in Ashbya gossypii |
author |
Ribeiro, Orquídea |
author_facet |
Ribeiro, Orquídea Wiebe, Marilyn Ilmén, Marja Domingues, Lucília Penttilä, Merja |
author_role |
author |
author2 |
Wiebe, Marilyn Ilmén, Marja Domingues, Lucília Penttilä, Merja |
author2_role |
author author author author |
dc.contributor.none.fl_str_mv |
Universidade do Minho |
dc.contributor.author.fl_str_mv |
Ribeiro, Orquídea Wiebe, Marilyn Ilmén, Marja Domingues, Lucília Penttilä, Merja |
dc.subject.por.fl_str_mv |
Ashbya gossypii Recombinant protein production Trichoderma reesei endoglucanase I Trichoderma reesei cellobiohydrolase I Cellulases heterologous expression Science & Technology |
topic |
Ashbya gossypii Recombinant protein production Trichoderma reesei endoglucanase I Trichoderma reesei cellobiohydrolase I Cellulases heterologous expression Science & Technology |
description |
To explore the potential of Ashbya gossypii as a host for the expression of recombinant proteins and to assess whether protein secretion would be more similar to the closely related Saccharomyces cerevisiae or to other filamentous fungi, endoglucanase I (EGI) and cellobiohydrolase I (CBHI) from the fungus Trichoderma reesei were successfully expressed in A. gossypii from plasmids containing the two micron sequences from S. cerevisiae, under the S. cerevisiae PGK1 promoter. The native signal sequences of EGI and CBHI were able to direct the secretion of EGI and CBHI into the culture medium in A. gossypii. Although CBHI activity was not detected using 4- methylumbelliferyl-β-D-lactoside as substrate, the protein was detected by Western blot using monoclonal antibodies. EGI activity was detectable, the specific activity being comparable to that produced by a similar EGI producing S. cerevisiae construct. More EGI was secreted than CBHI, or more active protein was produced. Partial characterization of CBHI and EGI expressed in A. gossypii revealed overglycosylation when compared with the native T. reesei proteins, but the glycosylation was less extensive than on cellulases expressed in S. cerevisiae. |
publishDate |
2010 |
dc.date.none.fl_str_mv |
2010 2010-01-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/1822/11325 |
url |
http://hdl.handle.net/1822/11325 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
"Applied Microbiology and Biotechnology". ISSN 0175-7598. 87:4 (2010) 1437-1446. 0175-7598 10.1007/s00253-010-2610-7 20422178 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Springer Verlag |
publisher.none.fl_str_mv |
Springer Verlag |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
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Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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RCAAP |
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RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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