page_title: Glutamine synthetase
section_title: Regulation and inhibition
hierarchical_section_title: Glutamine synthetase / Regulation and inhibition
caption_attribution_description: English: MSO as an inhibitor of glutamate
context_page_description: Glutamine synthetase is an enzyme that plays an essential role in the metabolism of nitrogen by catalyzing the condensation of glutamate and ammonia to form glutamine:
  Glutamate + ATP + NH₃ → Glutamine + ADP + phosphate
  Glutamine synthetase uses ammonia produced by nitrate reduction, amino acid degradation, and photorespiration. The amide group of glutamate is a nitrogen source for the synthesis of glutamine pathway metabolites.
  Other reactions may take place via GS. Competition between ammonium ion and water, their binding affinities, and the concentration of ammonium ion, influences glutamine synthesis and glutamine hydrolysis. Glutamine is formed if an ammonium ion attacks the acyl-phosphate intermediate, while glutamate is remade if water attacks the intermediate. Ammonium ion binds more strongly than water to GS due to electrostatic forces between a cation and a negatively charged pocket. Another possible reaction is upon NH₂OH binding to GS, rather than NH₄+, yields γ-glutamylhydroxamate.
context_section_description: Regulation of GS only occurs in prokaryotes. GS is subject to reversible covalent modification. Tyr³⁹⁷ of all 12 subunits can undergo adenylylation or deadenylylation by adenylyl transferase (AT), a bifunctional regulatory enzyme. Adenylylation is a post-translational modification involving the covalent attachment of AMP to a protein side chain. Each adenylylation requires an ATP and complete inhibition of GS requires 12 ATP. Deadenylylation by AT involves phosphorolytic removal of the Tyr-linked adenylyl groups as ADP. AT activity is influenced by the regulatory protein that is associated with it: PII, a 44-kD trimer. PII also undergoes post-translational modification by uridylyl transferase, thus PII has two forms. The state of PII dictates the activity of adenylyl transferase. If PII is not uridylylated, then it will take on the PIIA form. The AT:PIIA complex will deactivate GS by adenylylation. If PII is uridylylated, then it will take on the PIID form. The AT:PIID complex will activate GS by deadenylylation. The AT:PIIA and AT:PIID complexes are allosterically regulated in a reciprocal fashion by α-ketoglutarate (α-KG) and glutamine (Gln). Gln will activate AT:PIIA activity and inhibits AT:PIID, leading to adenylylation and subsequent deactivation of GS. Furthermore, Gln favors the conversion of PIID to PIIA. The effects of α-KG on the complexes are opposite. In the majority of gram-negative bacteria, GS can be modified by adenylylation (some cyanobacteria and green algae or exceptions).
  Inhibition of GS has largely focused on amino site ligands. Other inhibitors are the result of glutamine metabolism: tryptophan, histidine, carbamoyl phosphate, glucosamine-6-phosphate, cytidine triphosphate (CTP), and adenosine monophosphate (AMP). Other inhibitors/regulators are glycine and alanine. Alanine, glycine, and serine bind to the glutamate substrate site. GDP, AMP, ADP bind to the ATP site. L-serine, L-alanine, and glycine bind to the site for L-glutamate in unadenylated GS. The four amino acids bind to the site by their common atoms, “the main chain” of amino acids. Glutamate is another product of glutamine metabolism; however, glutamate is a substrate for GS inhibiting it to act as a regulator to GS.2 Each inhibitor can reduce the activity of the enzyme; once all final glutamine metabolites are bound to GS, the activity of GS is almost completely inhibited. Many inhibitory input signals allows for fine tuning of GS by reflecting nitrogen levels in the organism.
  Feedback regulation distinguishes the difference between two eukaryotic types of GS: brain and non-brain tissues. Non-brain GS responds to end-product feedback inhibition, while brain GS does not.  High concentrations of glutamine-dependent metabolites should inhibit GS activity, while low concentrations should activate GS activity.
  Inhibitors:
  Methionine sulfoximine (MSO): MSO is an inhibitor that binds to the glutamate site. Bound to GS, MSO is phosphorylated by ATP that results in an irreversible, non-covalent inhibition of GS. The S-isomer configuration is more inhibitory. Glutamate entry is blocked into the active site by a stabilization of the flexible loop in the active site by MSO.
  Phosphinothricin(PPT, Glufosinate): Phosphinothricin is an inhibitor that binds to the glutamate site. Glufosinate is used as an herbicide. Glufosinate treated plants die due to a buildup of ammonia and a cessation of photosynthesis.
  Many synthetic inhibitors are available today.
  Research on E. coli revealed that GS is regulated through gene expression. The gene that encodes the GS subunit is designated glnA. Transcription of glnA is dependent on NRI (a specific transcriptional enhancer). Active transcription occurs if NRI is in its phosphorylated form, designated NRI-P. Phosphorylation of NRI is catalyzed by NRII, a protein kinase. If NRII is complexed with PIIA then it will function as a phosphatase and NRI-P is converted back to NRI. In this case, transcription of glnA ceases.
  GS is subject to completely different regulatory mechanisms in cyanobacteria. Instead of the commo
caption: Methionine sulfoximine acting as an inhibitor to the glutamate binding site
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