Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation
Autor(a) principal: | |
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Data de Publicação: | 2023 |
Outros Autores: | , , , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | https://hdl.handle.net/1822/87235 |
Resumo: | Chitinases are widely studied enzymes that have already found widespread application. Their continued development and valorisation will be driven by the identification of new and improved variants and/or novel applications bringing benefits to industry and society. We previously identified a novel application for chitinases wherein the Candida albicans cell wall surface chitinase 3 (Cht3) was shown to have potential in vaccine applications as a subunit antigen against fungal infections. In the present study, this enzyme was investigated further, developing production and purification protocols, enriching our understanding of its properties, and advancing its application potential. Cht3 was heterologously expressed in Pichia pastoris and a 4-step purification protocol developed and optimised: this involves activated carbon treatment, hydrophobic interaction chromatography, ammonium sulphate precipitation, and gel filtration chromatography. The recombinant enzyme was shown to be mainly O-glycosylated and to retain the epitopes of the native protein. Functional studies showed it to be highly specific, displaying activity on chitin, chitosan, and chito-oligosaccharides larger than chitotriose only. Furthermore, it was shown to be a stable enzyme, exhibiting activity, and stability over broad pH and temperature ranges. This study represents an important step forward in our understanding of Cht3 and contributes to its development for application. |
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Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisationChitinaseEnzyme functionProtein productionProtein purificationVaccine antigenChitinases are widely studied enzymes that have already found widespread application. Their continued development and valorisation will be driven by the identification of new and improved variants and/or novel applications bringing benefits to industry and society. We previously identified a novel application for chitinases wherein the Candida albicans cell wall surface chitinase 3 (Cht3) was shown to have potential in vaccine applications as a subunit antigen against fungal infections. In the present study, this enzyme was investigated further, developing production and purification protocols, enriching our understanding of its properties, and advancing its application potential. Cht3 was heterologously expressed in Pichia pastoris and a 4-step purification protocol developed and optimised: this involves activated carbon treatment, hydrophobic interaction chromatography, ammonium sulphate precipitation, and gel filtration chromatography. The recombinant enzyme was shown to be mainly O-glycosylated and to retain the epitopes of the native protein. Functional studies showed it to be highly specific, displaying activity on chitin, chitosan, and chito-oligosaccharides larger than chitotriose only. Furthermore, it was shown to be a stable enzyme, exhibiting activity, and stability over broad pH and temperature ranges. This study represents an important step forward in our understanding of Cht3 and contributes to its development for application.Fundação para a Ciência e a Tecnologia (FCT), Grant/Award Numbers: UIDP/04050/2020, LA/P/0069/2020, SFRH/BD/133513/2017, COVID/BD/152169/2021, UI/BD/150872/2021WileyUniversidade do MinhoBarbosa, Augusto Alexandre CostaFerreira, DiogoPacheco, Maria InêsCasal, MargaridaDuarte, Henrique OliveiraGomes, CatarinaBarbosa, Ana Margarida MartinsTorrado, EgídioSampaio, PaulaCollins, Tony20232023-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttps://hdl.handle.net/1822/87235engCosta-Barbosa, A., Ferreira, D., Pacheco, M. I., Casal, M., Duarte, H. O., Gomes, C., Barbosa, A. M., Torrado, E., Sampaio, P., & Collins, T. (2023). Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation. Biotechnology Journal, 00, e2300219. https://doi.org/10.1002/biot.2023002191860-67681860-731410.1002/biot.20230021937876300https://onlinelibrary.wiley.com/doi/10.1002/biot.202300219info: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:RCAAP2024-02-24T01:23:00Zoai:repositorium.sdum.uminho.pt:1822/87235Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T22:53:58.725110Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
title |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
spellingShingle |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation Barbosa, Augusto Alexandre Costa Chitinase Enzyme function Protein production Protein purification Vaccine antigen |
title_short |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
title_full |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
title_fullStr |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
title_full_unstemmed |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
title_sort |
Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation |
author |
Barbosa, Augusto Alexandre Costa |
author_facet |
Barbosa, Augusto Alexandre Costa Ferreira, Diogo Pacheco, Maria Inês Casal, Margarida Duarte, Henrique Oliveira Gomes, Catarina Barbosa, Ana Margarida Martins Torrado, Egídio Sampaio, Paula Collins, Tony |
author_role |
author |
author2 |
Ferreira, Diogo Pacheco, Maria Inês Casal, Margarida Duarte, Henrique Oliveira Gomes, Catarina Barbosa, Ana Margarida Martins Torrado, Egídio Sampaio, Paula Collins, Tony |
author2_role |
author author author author author author author author author |
dc.contributor.none.fl_str_mv |
Universidade do Minho |
dc.contributor.author.fl_str_mv |
Barbosa, Augusto Alexandre Costa Ferreira, Diogo Pacheco, Maria Inês Casal, Margarida Duarte, Henrique Oliveira Gomes, Catarina Barbosa, Ana Margarida Martins Torrado, Egídio Sampaio, Paula Collins, Tony |
dc.subject.por.fl_str_mv |
Chitinase Enzyme function Protein production Protein purification Vaccine antigen |
topic |
Chitinase Enzyme function Protein production Protein purification Vaccine antigen |
description |
Chitinases are widely studied enzymes that have already found widespread application. Their continued development and valorisation will be driven by the identification of new and improved variants and/or novel applications bringing benefits to industry and society. We previously identified a novel application for chitinases wherein the Candida albicans cell wall surface chitinase 3 (Cht3) was shown to have potential in vaccine applications as a subunit antigen against fungal infections. In the present study, this enzyme was investigated further, developing production and purification protocols, enriching our understanding of its properties, and advancing its application potential. Cht3 was heterologously expressed in Pichia pastoris and a 4-step purification protocol developed and optimised: this involves activated carbon treatment, hydrophobic interaction chromatography, ammonium sulphate precipitation, and gel filtration chromatography. The recombinant enzyme was shown to be mainly O-glycosylated and to retain the epitopes of the native protein. Functional studies showed it to be highly specific, displaying activity on chitin, chitosan, and chito-oligosaccharides larger than chitotriose only. Furthermore, it was shown to be a stable enzyme, exhibiting activity, and stability over broad pH and temperature ranges. This study represents an important step forward in our understanding of Cht3 and contributes to its development for application. |
publishDate |
2023 |
dc.date.none.fl_str_mv |
2023 2023-01-01T00:00:00Z |
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 |
https://hdl.handle.net/1822/87235 |
url |
https://hdl.handle.net/1822/87235 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Costa-Barbosa, A., Ferreira, D., Pacheco, M. I., Casal, M., Duarte, H. O., Gomes, C., Barbosa, A. M., Torrado, E., Sampaio, P., & Collins, T. (2023). Candida albicans chitinase 3 with potential as a vaccine antigen: production, purification, and characterisation. Biotechnology Journal, 00, e2300219. https://doi.org/10.1002/biot.202300219 1860-6768 1860-7314 10.1002/biot.202300219 37876300 https://onlinelibrary.wiley.com/doi/10.1002/biot.202300219 |
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 |
Wiley |
publisher.none.fl_str_mv |
Wiley |
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 |
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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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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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1799135138067513344 |