title: The three-dimensional structure of an eukaryotic glutamine synthetase: functional implications of its oligomeric structure.
abstract: The structure of the prokaryotic glutamine synthetases type I (GS-I), key enzymes in nitrogen metabolism, was determined several years ago by X-ray diffraction, and consists of a double hexameric ring. The structure of the eukaryotic GS from the plant Phaseolus vulgaris (Glutamine synthetase type II; GS-II) has now been determined at low-resolution using electron microscopy and image processing, and consists of an octamer composed of two tetramers placed back-to-back and rotated 90 degrees with respect to each other. The oligomeric structure possesses a twofold symmetry, very suggestive of each tetramer being composed of two dimers. This is reinforced by the fact that dimers are isolated as a stable albeit non-functional species during the purification procedure. Given the fact that the active site of all types of GS is formed by highly conserved residues located in the interface of two interacting monomers, the geometry of the reconstructed tetramer suggests that it only contains two functional active sites, i.e., an active site per dimer. This is supported by biochemical data, which reveal that while the octamer binds eight ATP molecules, it only binds four molecules of the transition state analogue and GS inhibitor methionine-(S)-sulfoximine-P (MetSox-P). All this suggests for the GS-II enzyme an oligomeric structure containing four active sites and four possible regulatory sites, which might point to a complex regulatory behavior.
authors.affiliation: Centro Nacional de Biotecnología, C.S.I.C. Campus de la Universidad Autónoma de Madrid, Darwin 3, 28049 Madrid, Spain.
authors.forename: Oscar
authors.initials: O
authors.lastname: Llorca
authors.forename: Marco
authors.initials: M
authors.lastname: Betti
authors.forename: José M
authors.initials: JM
authors.lastname: González
authors.forename: Alfonso
authors.initials: A
authors.lastname: Valencia
authors.forename: Antonio J
authors.initials: AJ
authors.lastname: Márquez
authors.forename: José M
authors.initials: JM
authors.lastname: Valpuesta
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D002846:Chromatography, Affinity; D019281:Dimerization; D005974:Glutamate-Ammonia Ligase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007091:Image Processing, Computer-Assisted / Q000379:methods; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008717:Methionine Sulfoximine / Q000378:metabolism; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D027805:Phaseolus / Q000201:enzymology*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D008717:Methionine Sulfoximine; C083818:glutamine synthetase 2; D005974:Glutamate-Ammonia Ligase