Repair of Deaminated Base Damage by Schizosaccharomyces Pombe Thymine DNA Glycosylase

DNA Repair (Amst). 2008 Dec 1;7(12):1962-72. doi: 10.1016/j.dnarep.2008.08.006. Epub 2008 Sep 25.

Abstract

Thymine DNA glycosylases (TDG) in eukaryotic organisms are known for their double-stranded glycosylase activity on guanine/uracil (G/U) base pairs. Schizosaccharomyces pombe (Spo) TDG is a member of the MUG/TDG family that belongs to a uracil DNA glycosylase superfamily. This work investigates the DNA repair activity of Spo TDG on all four deaminated bases: xanthine (X) and oxanine (O) from guanine, hypoxanthine (I) from adenine, and uracil from cytosine. Unexpectedly, Spo TDG exhibits glycosylase activity on all deaminated bases in both double-stranded and single-stranded DNA in the descending order of X>I>U>>O. In comparison, human TDG only excises deaminated bases from G/U and, to a much lower extent, A/U and G/I base pairs. Amino acid substitutions in motifs 1 and 2 of Spo TDG show a significant impact on deaminated base repair activity. The overall mutational effects are characterized by a loss of glycosylase activity on oxanine in all five mutants. L157I in motif 1 and G288M in motif 2 retain xanthine DNA glycosylase (XDG) activity but reduce excision of hypoxanthine and uracil, in particular in C/I, single-stranded hypoxanthine (ss-I), A/U, and single-stranded uracil (ss-U). A proline substitution at I289 in motif 2 causes a significant reduction in XDG activity and a loss of activity on C/I, ss-I, A/U, C/U, G/U, and ss-U. S291G only retains reduced activity on T/I and G/I base pairs. S163A can still excise hypoxanthine and uracil in mismatched base pairs but loses XDG activity, making it the closest mutant, functionally, to human TDG. The relationship among amino acid substitutions, binding affinity and base recognition is discussed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • DNA Damage*
  • DNA Repair*
  • DNA, Fungal / metabolism*
  • DNA, Single-Stranded / genetics
  • Electrophoretic Mobility Shift Assay
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Purine Nucleosides / chemistry
  • Purine Nucleosides / metabolism
  • Schizosaccharomyces / enzymology*
  • Schizosaccharomyces / genetics
  • Sequence Homology, Amino Acid
  • Sequence Homology, Nucleic Acid
  • Thymine DNA Glycosylase / physiology*
  • Uracil / chemistry*
  • Uracil / metabolism
  • Xanthine / chemistry
  • Xanthine / metabolism

Substances

  • DNA, Fungal
  • DNA, Single-Stranded
  • Purine Nucleosides
  • oxanine
  • Xanthine
  • Uracil
  • Thymine DNA Glycosylase