Identification of tyrosine as a functional residue in the active site of Escherichia coli dUTPase

Biochim Biophys Acta. 1994 Mar 16;1205(1):146-50. doi: 10.1016/0167-4838(94)90103-1.

Abstract

dUTP nucleotidohydrolase (dUTPase, EC 3.6.1.23) from E. coli contains a total of six tyrosine residues per trimer. About half of them were found to be susceptible to acetylation with N-acetylimidazole or to nitration with tetranitromethane with concomitant loss of activity. Deacetylation with N-hydroxylamine leads to full reactivation. Inhibitory products of dUTP hydrolysis, i.e., dUMP and inorganic pyrophosphate together with the cofactor Mg2+ protect significantly against inactivation and chemical modification. In the Cu(2+)-dUTPase complex, charge transfer from Cu2+ to the tyrosinate anion was perturbed by the presence of the substrate dUTP. These results, together with the occurrence of one tyrosine residue in a strictly conserved sequence motif suggest the critical importance of this residue for the function of the enzyme.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylation
  • Amino Acid Sequence
  • Binding Sites
  • Escherichia coli / enzymology*
  • Imidazoles / pharmacology
  • Molecular Sequence Data
  • Pyrophosphatases / antagonists & inhibitors
  • Pyrophosphatases / chemistry*
  • Pyrophosphatases / isolation & purification
  • Tetranitromethane / pharmacology
  • Tyrosine / analysis*

Substances

  • Imidazoles
  • Tyrosine
  • Pyrophosphatases
  • dUTP pyrophosphatase
  • Tetranitromethane
  • N-acetylimidazole