No abstract

In vertebrates, membrane-bound ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) on the cell surface are responsible for signal conversion and termination in purinergic signaling by extracellular nucleotides. Here we present apo and complex structures of the rat NTPDase2 extracellular domain and Legionella pneumophila NTPDase1, including a high-resolution structure with a transition-state analog. Comparison of ATP and ADP binding modes shows how NTPDases engage the same catalytic site for hydrolysis of nucleoside triphosphates and diphosphates. We find that this dual specificity is achieved at the expense of base specificity. Structural and mutational studies indicate that a conserved active-site water is replaced by the phosphate product immediately after phosphoryl transfer. Partial base specificity for purines in LpNTPDase1 is based on a different intersubunit base binding site for pyrimidine bases. A comparison of the bacterial enzyme in six independent crystal forms shows that NTPDases can undergo a domain closure motion of at least 17°.

Background: Disulfide-regulated NTPDases from T. gondii are related to the virulence of the parasite. Results: Crystal structures of the active and inactive state were determined. Conclusion: The 258 -268 disulfide bridge acts like a clamp, which upon reduction allows concerted motions of the subunits and domains of the tetrameric enzyme. Significance: First molecular model of the disulfide-regulated activation mode.

Nucleoside triphosphate diphosphohydrolases (NTPDases) are secreted or membrane-bound ectonucleotidases that hydrolyze the anhydride bonds of nucleoside triphosphates and nucleoside diphosphates. Mammalian cell-surface NTPDase enzymes are inhibited by various polyoxometallates. Here, the structures of NTPDase1 from the bacterium Legionella pneumophila (LpNTPDase1) in complex with the dodecatungstate POM-1, decavanadate and octamolybdate/heptamolybdate are described. The metal clusters are bound at different sites but always in a highly ordered fashion via electrostatic interactions and hydrogen bonds. For octamolybdate, covalent interactions after oxygen ligand exchange by a serine and histidine side chain are also observed. The potential inhibitory mechanism and the use of the metal clusters as phasing tools for new NTPDase structures are discussed. The binding mode of a tartrate ion at the catalytic centre suggests novel strategies for the structure-based design of NTPDase inhibitors, and the observation of the enzyme in an intermediate open state contributes to our understanding of NTPDase enzyme dynamics.

La première solution, qui parait peut-être extrême. a cependant déjà été avancée par des États. Voir, par exemple, la déclaration mexicaine lors de la guerre civile espagnole, Journal officiel de fa Société des Nations, mars 1937, 264. 21. L'expression est de nous. Plusieurs auteurs qualifient cette doctrine majoritaire de "théorie de non-intervention": voir ex. OGLESBY, op. cit., supra, note 6. Cette désignation est douteuse. En effet, pour plusieurs, "non-intervention" a une autre acceptioncelle d'une interdiction symétrique d'aider toutes les parties au conflit. N'était-ce pas Talleyrand qui dit: "Non-intervention est un terme plutôt mystérieux, et qui signifie intervention"? C'est le désir d'éviter cette confusion qui a dicté notre choix terminologique.