A New PII Protein Structure Identifies the 2-Oxoglutarate Binding Site

PII proteins of bacteria, archaea, and plants regulate many facets of nitrogen metabolism. They do so by interacting with their target proteins, which can be enzymes, transcription factors, or membrane proteins. A key feature of the ability of PII proteins to sense cellular nitrogen status and to interact accordingly with their targets is their binding of the key metabolic intermediate 2-oxoglutarate (2-OG). However, the binding site of this ligand within PII proteins has been controversial. We have now solved the X-ray structure, at 1.4 Å resolution, of the Azospirillum brasilense PII protein GlnZ complexed with MgATP and 2-OG. This structure is in excellent agreement with previous biochemical data on 2-OG binding to a variety of PII proteins and shows that 2-oxglutarate binds within the cleft formed between neighboring subunits of the homotrimer. The 2-oxo acid moiety of bound 2-OG ligates the bound Mg2+ together with three phosphate oxygens of ATP and the side chain of the T-loop residue Gln39. Our structure is in stark contrast to an earlier structure of the Methanococcus jannaschii GlnK1 protein in which the authors reported 2-OG binding to the T-loop of that PII protein. In the light of our new structure, three families of T-loop conformations, each associated with a distinct effector binding mode and characterized by a different interaction partner of the ammonium group of the conserved residue Lys58, emerge as a common structural basis for effector signal output by PII proteins.