3F4F

Crystal structure of dUT1p, a dUTPase from Saccharomyces cerevisiae

3F4F の概要

• エントリーDOI: 10.2210/pdb3f4f/pdb
• 分子名称: Deoxyuridine 5'-triphosphate nucleotidohydrolase, 2'-DEOXYURIDINE 5'-MONOPHOSPHATE, SODIUM ION, ... (6 entities in total)
• 機能のキーワード: trimer, beta barrel, ump, product complex, dutp pyrophosphatase, ditp, hydrolase, nucleotide metabolism, phosphoprotein
• 由来する生物種: Saccharomyces cerevisiae
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 53767.84

構造登録者

Singer, A.U.,Evdokimova, E.,Kudritska, M.,Edwards, A.M.,Yakunin, A.F.,Savchenko, A. (登録日: 2008-10-31, 公開日: 2008-11-11, 最終更新日: 2023-09-06)

主引用文献

Tchigvintsev, A.,Singer, A.U.,Flick, R.,Petit, P.,Brown, G.,Evdokimova, E.,Savchenko, A.,Yakunin, A.F.
Structure and activity of the Saccharomyces cerevisiae dUTP pyrophosphatase DUT1, an essential housekeeping enzyme.
Biochem.J., 437:243-253, 2011

PubMed Abstract: 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 Å resolution (1 Å=0.1 nm) in an apo state and in complex with the non-hydrolysable substrate α,β-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.

PubMed: 21548881
DOI: 10.1042/BJ20110304

実験手法

X-RAY DIFFRACTION (2 Å)