3p48

Structure of the yeast dUTPase DUT1 in complex with dUMPNPP

spinqualitylabelsall models 3p48, resolution 1.67Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 3p48 is a 3 chain structure with sequence from Saccharomyces cerevisiae. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 1.67Å Ligands: DUP, MG Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function DUT_YEAST This enzyme is involved in nucleotide metabolism: it produces dUMP, the immediate precursor of thymidine nucleotides and it decreases the intracellular concentration of dUTP so that uracil cannot be incorporated into DNA. Publication Abstract from PubMed Genomes of all free-living organisms encode the enzyme dUTPase (dUTP pyrophosphatase), which plays a key role in preventing uracil incorporation into DNA. In the present paper, we describe the biochemical and structural characterization of DUT1 (Saccharomyces cerevisiae dUTPase). The hydrolysis of dUTP by DUT1 was strictly dependent on a bivalent metal cation with significant activity observed in the presence of Mg2+, Co2+, Mn2+, Ni2+ or Zn2+. In addition, DUT1 showed a significant activity against another potentially mutagenic nucleotide: dITP. With both substrates, DUT1 demonstrated a sigmoidal saturation curve, suggesting a positive co-operativity between the subunits. The crystal structure of DUT1 was solved at 2 A resolution (1 A=0.1 nm) in an apo state and in complex with the non-hydrolysable substrate alpha,beta-imido dUTP or dUMP product. Alanine-replacement mutagenesis of the active-site residues revealed seven residues important for activity including the conserved triad Asp87/Arg137/Asp85. The Y88A mutant protein was equally active against both dUTP and UTP, indicating that this conserved tyrosine residue is responsible for discrimination against ribonucleotides. The structure of DUT1 and site-directed mutagenesis support a role of the conserved Phe142 in the interaction with the uracil base. Our work provides further insight into the molecular mechanisms of substrate selectivity and catalysis of dUTPases. Structure and activity of the Saccharomyces cerevisiae dUTP pyrophosphatase DUT1, an essential housekeeping enzyme.,Tchigvintsev A, Singer AU, Flick R, Petit P, Brown G, Evdokimova E, Savchenko A, Yakunin AF Biochem J. 2011 Jul 15;437(2):243-53. PMID:21548881 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See Also DUTPase 3D structures

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