title: Chemical and thermal influence of the [4Fe-4S]2+ cluster of A/G-specific adenine glycosylase from Corynebacterium pseudotuberculosis.
abstract: The gram-positive bacteria Corynebacterium pseudotuberculosis, the causative agent of caseous lymphadenitis in livestock significantly reduces productivity and often causes death. The adenine/guanine-specific DNA glycosylase (MutY) prevents mutations in the DNA of the pathogen and a unique feature of the MutY protein family is the [4Fe-4S]2+ cluster that interlinks two protein subdomains. MutY from C. pseudotuberculosis was expressed in E. coli and purified, the CD experiments indicate a high content of α-helices and random coiled secondary structure and a typical near-UV CD fingerprint for the [4Fe-4S]2+ cluster. EDTA and copper sulfate possess a strong destabilizing effect on the [4Fe-4S]2+ cluster. UV-vis and fluorescence spectroscopy results demonstrate that between pH3.0 and 4.0 the integrity of the [4Fe-4S]2+ cluster is destroyed. To investigate the thermal stability of the protein differential scanning calorimetry and fluorescence spectroscopy were used and the Tm was determined to be 45°C. The analysis presented provides information concerning the protein stability under different physio-chemical conditions.
authors.affiliation: Multiuser Center for Biomolecular Innovation, Departament of Physics, Universidade Estadual Paulista (UNESP), São Jose do Rio Preto, SP 15054-000, Brazil.
authors.forename: Raphael J
authors.initials: RJ
authors.lastname: Eberle
authors.affiliation: Multiuser Center for Biomolecular Innovation, Departament of Physics, Universidade Estadual Paulista (UNESP), São Jose do Rio Preto, SP 15054-000, Brazil.
authors.forename: Monika A
authors.initials: MA
authors.lastname: Coronado
authors.affiliation: Multiuser Center for Biomolecular Innovation, Departament of Physics, Universidade Estadual Paulista (UNESP), São Jose do Rio Preto, SP 15054-000, Brazil.
authors.forename: Icaro P
authors.initials: IP
authors.lastname: Caruso
authors.affiliation: Laboratory of Molecular Biology, Federal University of São João Del-Rei (CCO), Av. Sebastião Gonçalves Coelho, 400, Divinópolis, MG 35501-296, Brazil.
authors.forename: Débora O
authors.initials: DO
authors.lastname: Lopes
authors.affiliation: Laboratório de Genética Celular e Molecular, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627-Pampulha, CP 486, CEP 31, Belo Horizonte, MG 270-901, Brazil.
authors.forename: Anderson
authors.initials: A
authors.lastname: Miyoshi
authors.affiliation: Laboratório de Genética Celular e Molecular, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627-Pampulha, CP 486, CEP 31, Belo Horizonte, MG 270-901, Brazil.
authors.forename: Vasco
authors.initials: V
authors.lastname: Azevedo
authors.affiliation: Multiuser Center for Biomolecular Innovation, Departament of Physics, Universidade Estadual Paulista (UNESP), São Jose do Rio Preto, SP 15054-000, Brazil. Electronic address: arni@sjrp.unesp.br.
authors.forename: Raghuvir K
authors.initials: RK
authors.lastname: Arni
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002942:Circular Dichroism; D019327:Copper Sulfate / Q000737:chemistry; D016925:Corynebacterium pseudotuberculosis / Q000201:enzymology* / Q000235:genetics; D045647:DNA Glycosylases / Q000737:chemistry* / Q000235:genetics; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; D045647:DNA Glycosylases; C061738:mutY adenine glycosylase; D019327:Copper Sulfate
keywords: C. pseudotuberculosis; DNA repair; MutY; Secondary and tertiary structure; Spectroscopic method; [4Fe–4S](2+) cluster