Structural insights into the catalytic mechanism of phosphate ester hydrolysis by dUTPase

Orsolya Barabás, Veronika Pongrácz, Júlia Kovári, Matthias Wilmanns, Beáta G. Vértessy

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66 Citations (Scopus)

Abstract

dUTPase is essential to keep uracil out of DNA. Crystal structures of substrate (dUTP and α,β-imino-dUTP) and product complexes of wild type and mutant dUTPases were determined to reveal how an enzyme responsible for DNA integrity functions. A kinetic analysis of wild type and mutant dUTPases was performed to obtain relevant mechanistic information in solution. Substrate hydrolysis is shown to be initiated via in-line nucleophile attack of a water molecule oriented by an activating conserved aspartate residue. Substrate binding in a catalytically competent conformation is achieved by (i) multiple interactions of the triphosphate moiety with catalysis-assisting Mg 2+, (ii) a concerted motion of residues from three conserved enzyme motifs as compared with the apoenzyme, and (iii) an intricate hydrogen-bonding network that includes several water molecules in the active site. Results provide an understanding for the catalytic role of conserved residues in dUTPases.

Original languageEnglish
Pages (from-to)42907-42915
Number of pages9
JournalJournal of Biological Chemistry
Volume279
Issue number41
DOIs
Publication statusPublished - Oct 8 2004

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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