title: Structure and enzymatic mechanism of a moonlighting dUTPase.
abstract: Genome integrity requires well controlled cellular pools of nucleotides. dUTPases are responsible for regulating cellular dUTP levels and providing dUMP for dTTP biosynthesis. In Staphylococcus, phage dUTPases are also suggested to be involved in a moonlighting function regulating the expression of pathogenicity-island genes. Staphylococcal phage trimeric dUTPase sequences include a specific insertion that is not found in other organisms. Here, a 2.1 Å resolution three-dimensional structure of a ϕ11 phage dUTPase trimer with complete localization of the phage-specific insert, which folds into a small β-pleated mini-domain reaching out from the dUTPase core surface, is presented. The insert mini-domains jointly coordinate a single Mg2+ ion per trimer at the entrance to the threefold inner channel. Structural results provide an explanation for the role of Asp95, which is suggested to have functional significance in the moonlighting activity, as the metal-ion-coordinating moiety potentially involved in correct positioning of the insert. Enzyme-kinetics studies of wild-type and mutant constructs show that the insert has no major role in dUTP binding or cleavage and provide a description of the elementary steps (fast binding of substrate and release of products). In conclusion, the structural and kinetic data allow insights into both the phage-specific characteristics and the generally conserved traits of ϕ11 phage dUTPase.
authors: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, 29 Karolina Street, 1113 Budapest, Hungary.
authors: Ibolya
authors: I
authors: Leveles
authors: Veronika
authors: V
authors: Németh
authors: Judit E
authors: JE
authors: Szabó
authors: Veronika
authors: V
authors: Harmat
authors: Kinga
authors: K
authors: Nyíri
authors: Ábris Ádám
authors: ÁÁ
authors: Bendes
authors: Veronika
authors: V
authors: Papp-Kádár
authors: Imre
authors: I
authors: Zagyva
authors: Gergely
authors: G
authors: Róna
authors: Olivér
authors: O
authors: Ozohanics
authors: Károly
authors: K
authors: Vékey
authors: Judit
authors: J
authors: Tóth
authors: Beáta G
authors: BG
authors: Vértessy
mesh_terms: D000595:Amino Acid Sequence; D002413:Cations, Divalent / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D011755:Pyrophosphatases / Q000737:chemistry* / Q000378:metabolism*; D016415:Sequence Alignment; D013204:Staphylococcus Phages / Q000737:chemistry / Q000201:enzymology*; D013211:Staphylococcus aureus / Q000821:virology
chemical_list: D002413:Cations, Divalent; D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase; D008274:Magnesium
keywords: dUTPases; moonlighting; ϕ11 phage dUTPase