Functional and structural studies of a mini-ferritin protein

Detalhes bibliográficos
Autor(a) principal: Macedo, Joana Tavares
Data de Publicação: 2014
Tipo de documento: Dissertação
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10362/13301
Resumo: Dissertação para obtenção do Grau de Mestre em Bioquímica
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spelling Functional and structural studies of a mini-ferritin proteinDpsX-ray crystallographyIron storageDNA bindingFerroxidase centerCopper bindingDissertação para obtenção do Grau de Mestre em BioquímicaDNA-binding protein from starved cells (Dps) are mini-ferritins mainly expressed in bacteria during severe environmental stress. These proteins with a highly conserved structure provide wide protection to cells and function as iron-storage proteins. Some Dps can also bind DNA and the N-terminus has been suggested to be involved in this interaction, due to its positively charged residues. This thesis focused on the functional and structural features of recombinant Dps protein from Pseudomonas nautica 617 (P. nautica). To investigate the iron incorporation mechanism, iron uptake assays using H2O2 and O2 as co-substrate for iron oxidation were performed with Dps WT and F46G variant, using UVVisible spectroscopy. The results showed that Phe46, located close to the ferroxidase center (FOC), does not influence the amount of iron stored. Nevertheless, this residue affectes the iron core formation when O2 was used as co-substrate. Structural characterization of the incorporation of Fe(II) and Cu(II) was performed with X-ray crystallography. High resolution crystal structures of Apo-Dps, Dps incubated with 12 Fe(II)/Dps (12Fe-Dps) and Dps incubated with 12 Cu(II)/Dps (12Cu-Dps) were obtained. It was possible to observe the binding of the metals to the FOCs with different coordination geometry as well as geometrically different FOCs. Additionally Fe(II) and Cu(II) atoms were assigned in a position where hydrophilic pores can be created and serve as entry routes for the metals. For the characterization of the Dps-DNA interaction, electrophoretic mobility shift assays (EMSAs) were carried out with Dps WT and Q14E variant. The results showed that this protein can bind DNA, but the affinity for DNA significantly decreases in the presence of the negative charge in the N-terminus. In this sense, mutations in the N-terminus that may increase the affinity for DNA binding were produced by site-directed mutagenesis.Faculdade de Ciências e TecnologiaTavares, PedroPereira, AliceAndrade, SusanaRUNMacedo, Joana Tavares2014-10-17T14:41:58Z2014-092014-09-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisapplication/pdfhttp://hdl.handle.net/10362/13301enginfo: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:RCAAP2024-03-11T03:48:07Zoai:run.unl.pt:10362/13301Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:21:13.998760Repositório 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 Functional and structural studies of a mini-ferritin protein
title Functional and structural studies of a mini-ferritin protein
spellingShingle Functional and structural studies of a mini-ferritin protein
Macedo, Joana Tavares
Dps
X-ray crystallography
Iron storage
DNA binding
Ferroxidase center
Copper binding
title_short Functional and structural studies of a mini-ferritin protein
title_full Functional and structural studies of a mini-ferritin protein
title_fullStr Functional and structural studies of a mini-ferritin protein
title_full_unstemmed Functional and structural studies of a mini-ferritin protein
title_sort Functional and structural studies of a mini-ferritin protein
author Macedo, Joana Tavares
author_facet Macedo, Joana Tavares
author_role author
dc.contributor.none.fl_str_mv Tavares, Pedro
Pereira, Alice
Andrade, Susana
RUN
dc.contributor.author.fl_str_mv Macedo, Joana Tavares
dc.subject.por.fl_str_mv Dps
X-ray crystallography
Iron storage
DNA binding
Ferroxidase center
Copper binding
topic Dps
X-ray crystallography
Iron storage
DNA binding
Ferroxidase center
Copper binding
description Dissertação para obtenção do Grau de Mestre em Bioquímica
publishDate 2014
dc.date.none.fl_str_mv 2014-10-17T14:41:58Z
2014-09
2014-09-01T00:00:00Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/masterThesis
format masterThesis
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10362/13301
url http://hdl.handle.net/10362/13301
dc.language.iso.fl_str_mv eng
language eng
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 Faculdade de Ciências e Tecnologia
publisher.none.fl_str_mv Faculdade de Ciências e Tecnologia
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
instname_str Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
instacron_str RCAAP
institution RCAAP
reponame_str Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
collection Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
repository.name.fl_str_mv Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
repository.mail.fl_str_mv
_version_ 1799137853295296512

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