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2005 | 52 | 1 | 167-178
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Inhibition of DNA repair glycosylases by base analogs and tryptophan pyrolysate, Trp-P-1.

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DNA base analogs, 2,4,5,6-substituted pyrimidines and 2,6-substituted purines were tested as potential inhibitors of E. coli Fpg protein (formamidopyrimidine -DNA glycosylase). Three of the seventeen compounds tested revealed inhibitory properties. 2-Thioxanthine was the most efficient, inhibiting 50% of 2,6-diamino-4-hydroxy-5N-methyl-formamidopyrimidine (Fapy-7MeG) excision activity at 17.1 μM concentration. The measured Kgi was 4.44 ± 0.15 μM. Inhibition was observed only when the Fpg protein was first challenged to its substrate followed by the addition of the base analog, suggesting uncompetitive (catalytic) inhibition. For two other compounds, 2-thio- or 2-oxo-4,5,6-substituted pyrimidines, IC50 was only 343.3 ± 58.6 and 350 ± 24.4 μM, respectively. No change of the Fpg glycosylase activity was detected in the presence of Fapy-7MeG, up to 5 μM. We also investigated the effect of DNA structure modified by tryptophan pyrolysate (Trp-P-1) on the activity of base excision repair enzymes: Escherichia coli and human DNA glycosylases of oxidized (Fpg, Nth) and alkylated bases (TagA, AlkA, and ANPG), and for bacterial AP endonuclease (Xth protein). Trp-P-1, which changes the secondary DNA structure into non-B, non-Z most efficiently inhibited excision of alkylated bases by the AlkA glycosylase (IC50 = 1 μM). The ANPG, TagA, and Fpg proteins were also inhibited although to a lesser extent (IC50 = 76.5 μM, 96 μM, and 187.5 μM, respectively). Trp-P-1 also inhibited incision of DNA at abasic sites by the β-lyase activity of the Fpg and Nth proteins, and to a lesser extent by the Xth AP endonuclease. Thus, DNA conformation is critical for excision of damaged bases and incision of abasic sites by DNA repair enzymes.
Physical description
  • Department of Molecular Biology, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warszawa, Poland
  • Department of Molecular Biology, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warszawa, Poland
  • Department of Molecular Biology, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warszawa, Poland
  • Groupe "Reparation de l'ADN," CNRS Unite Mixte de Recherche 8126, Institut Gustave Roussy, Villejuif Cedex, France
  • Laboratory of Experimental Pharmacology, Medical Research Center, Polish Academy of Sciences, Warszawa, Poland
  • Department of Molecular Biology, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warszawa, Poland
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