title: The toxin-antitoxin proteins relBE2Spn of Streptococcus pneumoniae: characterization and association to their DNA target.
abstract: The chromosome of the pathogenic Gram-positive bacterium Streptococcus pneumoniae contains between six to 10 operons encoding toxin-antitoxin systems (TAS). TAS are widespread and redundant in bacteria and archaea and their role, albeit still obscure, may be related to important aspects of bacteria lifestyle like response to stress. One of the most abundant TAS is the relBE family, being present in the chromosome of many bacteria and archaea. Because of the high rates of morbility and mortality caused by S. pneumoniae, it has been interesting to gain knowledge on the pneumococcal TAS, among them the RelBE2Spn proteins. Here, we have analyzed the DNA binding capacity of the RelB2Spn antitoxin and the RelB2Spn-RelE2Spn proteins by band-shift assays. Thus, a DNA region encompassing the operator region of the proteins was identified. In addition, we have used analytical ultracentrifugation and native mass spectrometry to measure the oligomerization state of the antitoxin alone and the RelBE2Spn complex in solution bound or unbound to its DNA substrate. Using native mass spectrometry allowed us to unambiguously determine the stoichiometry of the RelB2Spn and of the RelBE2Spn complex alone or associated to its DNA target.
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
authors.forename: Inma
authors.initials: I
authors.lastname: Moreno-Córdoba
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Diago-Navarro
authors.forename: Arjan
authors.initials: A
authors.lastname: Barendregt
authors.forename: Albert J R
authors.initials: AJ
authors.lastname: Heck
authors.forename: Carlos
authors.initials: C
authors.lastname: Alfonso
authors.forename: Ramón
authors.initials: R
authors.lastname: Díaz-Orejas
authors.forename: Concha
authors.initials: C
authors.lastname: Nieto
authors.forename: Manuel
authors.initials: M
authors.lastname: Espinosa
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013296:Streptococcus pneumoniae / Q000737:chemistry*; D014461:Ultracentrifugation
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D004247:DNA