title: 2-Oxoglutarate levels control adenosine nucleotide binding by Herbaspirillum seropedicae PII proteins.
abstract: Nitrogen metabolism in Proteobacteria is controlled by the Ntr system, in which PII proteins play a pivotal role, controlling the activity of target proteins in response to the metabolic state of the cell. Characterization of the binding of molecular effectors to these proteins can provide information about their regulation. Here, the binding of ATP, ADP and 2-oxoglutarate (2-OG) to the Herbaspirillum seropedicae PII proteins, GlnB and GlnK, was characterized using isothermal titration calorimetry. Results show that these proteins can bind three molecules of ATP, ADP and 2-OG with homotropic negative cooperativity, and 2-OG binding stabilizes the binding of ATP. Results also show that the affinity of uridylylated forms of GlnB and GlnK for nucleotides is significantly lower than that of the nonuridylylated proteins. Furthermore, fluctuations in the intracellular concentration of 2-OG in response to nitrogen availability are shown. Results suggest that under nitrogen-limiting conditions, PII proteins tend to bind ATP and 2-OG. By contrast, after an ammonium shock, a decrease in the 2-OG concentration is observed causing a decrease in the affinity of PII proteins for ATP. This phenomenon may facilitate the exchange of ATP for ADP on the ligand-binding pocket of PII proteins, thus it is likely that under low ammonium, low 2-OG levels would favor the ADP-bound state.
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Marco A S
authors.initials: MA
authors.lastname: Oliveira
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Edileusa C M
authors.initials: EC
authors.lastname: Gerhardt
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Luciano F
authors.initials: LF
authors.lastname: Huergo
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Emanuel M
authors.initials: EM
authors.lastname: Souza
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Fábio O
authors.initials: FO
authors.lastname: Pedrosa
authors.affiliation: Department of Biochemistry and Molecular Biology, Universidade Federal do Parana, Curitiba, Brazil.
authors.forename: Leda S
authors.initials: LS
authors.lastname: Chubatsu
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002151:Calorimetry; D005974:Glutamate-Ammonia Ligase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D042803:Herbaspirillum / Q000201:enzymology* / Q000502:physiology; D007656:Ketoglutaric Acids / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D009586:Nitrogen Fixation; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051922:PII Nitrogen Regulatory Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013312:Stress, Physiological; D017186:Titrimetry
chemical_list: D001426:Bacterial Proteins; D007656:Ketoglutaric Acids; D008024:Ligands; D051922:PII Nitrogen Regulatory Proteins; D011994:Recombinant Proteins; C404143:PIID regulatory protein, Bacteria; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D009713:Nucleotidyltransferases; D005974:Glutamate-Ammonia Ligase
keywords: 2-oxoglutarate signaling; Herbaspirillum seropedicae; Ntr system; PII proteins; nitrogen metabolism