Substrate specificity of Schizosaccharomyces pombe Nth protein for products of oxidative DNA damage.

Karahalil B., Roldan-Arjona T., Dizdaroglu M.

Biochemistry, vol.37, no.2, pp.590-5, 1998 (SCI-Expanded) identifier identifier identifier

  • Publication Type: Article / Article
  • Volume: 37 Issue: 2
  • Publication Date: 1998
  • Doi Number: 10.1021/bi971660s
  • Journal Name: Biochemistry
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Page Numbers: pp.590-5
  • Gazi University Affiliated: Yes


A gene from Schizosaccharomyces pombe, which encodes a protein with a strong sequence similarity to the Nth protein of Escherichia: coli, has recently been identified [Roldan-Arjona, T., Anselmino, C., and Lindahl, T. (1996) Nucleic Acids Res. 24, 3307-3312]. The functional analysis of this eukaryotic enzyme indicated that it is a homologue of E. call Nth protein. The gene has been subcloned and the protein (Nth-Spo) purified to apparent homogeneity. We investigated the substrate specificity of this eukaryotic enzyme for modified bases in oxidatively damaged DNA, using the technique of gas chromatography/isotope-dilution mass spectrometry (GC/IDMS). DNA substrates containing up to 17 types of modified bases were prepared by gamma-irradiation or by treatment with. H2O2 in the presence of Fe(III)-EDTA or Cu(II). The results revealed an efficient excision of five pyrimidine-derived lesions, 5-hydroxycytosine, thymine glycol, 5-hydroxy-6-hydrothymine: 5,6-dihydroxycytosine, and 5-hydroxyuracil. None of the other pyrimidine or purine lesions was excised. Excision was measured as a function of enzyme concentration, time, substrate concentration, and temperature. Kinetic constants were determined. Although some DNA base lesions removed by Nth-Spo protein were similar to those previously described for E. coti Nth protein, differences between substrate specificities of these two enzymes were noted.