title: Purification of the Campylobacter jejuni Dps protein assisted by its high melting temperature.
abstract: Dps proteins (DNA binding protein from starved cell) form a distinct group within the ferritin superfamily. All Dps members are composed of 12 identical subunits that assemble into a conserved spherical protein shell. Dps oxidize Fe(2+) in a conserved ferroxidase center located at the interface between monomers, the product of the reaction Fe(3+), is then stored inside the protein shell in the form of non-reactive insoluble Fe2O3. The Campylobacter jejuni Dps (CjDps) has been reported to play a plethora of functions, such as DNA binding and protection, iron storage, survival in response to hydrogen peroxide and sulfatide binding. CjDps is also important during biofilm formation and caecal colonization in poultry. In order to facilitate in vitro characterisation of CjDps, it is important to have a simple and reproducible protocol for protein purification. Here we report an observation that CjDps has an unusual high melting temperature. We exploited this property for protein purification by introducing a thermal treatment step which allowed achieving homogeneity by using only two chromatographic steps. Gel filtration chromatography, circular dichroism, mass spectrometry, DNA-binding and iron oxidation analysis confirmed that the CjDps structure and function were unaffected.
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Heloisa B S
authors.initials: HB
authors.lastname: Sanchuki
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Glaucio
authors.initials: G
authors.lastname: Valdameri
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Vivian R
authors.initials: VR
authors.lastname: Moure
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Marco A
authors.initials: MA
authors.lastname: Oliveira
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Fábio O
authors.initials: FO
authors.lastname: Pedrosa
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil.
authors.forename: Emanuel M
authors.initials: EM
authors.lastname: Souza
authors.affiliation: Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, Australia.
authors.forename: Victoria
authors.initials: V
authors.lastname: Korolik
authors.affiliation: Instituto Nacional de Ciência e Tecnologia da Fixação Biológica de Nitrogênio, Departamento de Bioquímica e Biologia Molecular, UFPR Curitiba, PR, Brazil. Electronic address: Huergo@ufpr.br.
authors.forename: Luciano F
authors.initials: LF
authors.lastname: Huergo
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D016123:Campylobacter jejuni / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D006358:Hot Temperature
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C083911:DPS protein, Bacteria
keywords: DNA binding; Dps protein; Heat treatment; Melting temperature