title: Crystal structure of the ATPPase subunit and its substrate-dependent association with the GATase subunit: a novel regulatory mechanism for a two-subunit-type GMP synthetase from Pyrococcus horikoshii OT3.
abstract: Guanosine 5'-monophosphate synthetase(s) (GMPS) catalyzes the final step of the de novo synthetic pathway of purine nucleotides. GMPS consists of two functional units that are present as domains or subunits: glutamine amidotransferase (GATase) and ATP pyrophosphatase (ATPPase). GATase hydrolyzes glutamine to yield glutamate and ammonia, while ATPPase utilizes ammonia to convert adenyl xanthosine 5'-monophosphate (adenyl-XMP) into guanosine 5'-monophosphate. Here we report the crystal structure of PH-ATPPase (the ATPPase subunit of the two-subunit-type GMPS from the hyperthermophilic archaeon Pyrococcus horikoshii OT3). PH-ATPPase consists of two domains (N-domain and C-domain) and exists as a homodimer in the crystal and in solution. The N-domain contains an ATP-binding platform called P-loop, whereas the C-domain contains the xanthosine 5'-monophosphate (XMP)-binding site and also contributes to homodimerization. We have also demonstrated that PH-GATase (the glutamine amidotransferase subunit of the two-subunit-type GMPS from the hyperthermophilic archaeon P. horikoshii OT3) alone is inactive, and that all substrates of PH-ATPPase except for ammonia (Mg(2+), ATP and XMP) are required to stabilize the active complex of PH-ATPPase and PH-GATase subunits.
authors.affiliation: Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
authors.forename: Shintaro
authors.initials: S
authors.lastname: Maruoka
authors.forename: Shoichiro
authors.initials: S
authors.lastname: Horita
authors.forename: Woo Cheol
authors.initials: WC
authors.lastname: Lee
authors.forename: Koji
authors.initials: K
authors.lastname: Nagata
authors.forename: Masaru
authors.initials: M
authors.lastname: Tanokura
mesh_terms: D000582:Amidophosphoribosyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000641:Ammonia / Q000494:pharmacology; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019731:Carbon-Nitrogen Ligases; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D043902:Pyrococcus horikoshii / Q000201:enzymology* / Q000235:genetics; D011755:Pyrophosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D040681:Structural Homology, Protein; D013379:Substrate Specificity
chemical_list: D019843:Archaeal Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D000641:Ammonia; D000582:Amidophosphoribosyltransferase; D011755:Pyrophosphatases; C019757:ATP pyrophosphohydrolase; D019731:Carbon-Nitrogen Ligases; D019733:Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor; C020828:GMP synthase (glutamine-hydrolyzing)