Processing of Thymine Glycols, Urea Residues and AP Sites in Escherichia Coli

  • Michael F. Laspia
  • Susan S. Wallace


In order to examine the biological consequences of oxidative DNA damage, we have selectively introduced damages of interest into phage transfecting DNA so that we can examine the repair of these lesions in vivo. Thymine glycol is a relatively stable ring saturation product of thymine that is formed as a consequence of hydroxyl radical attack on the 5,6 double bond. Thymine glycol is found in DNA X-irradiated in vitro (1,2) and in vivo (3,4) and appears to be formed as a consequence of oxidative stress (5). Thymine glycol is a replicative block to DNA polymerases in vitro (6–9), however, it retains pairing capacity since an A is inserted opposite the lesion before synthesis is arrested (10). Urea residues are fragmentation products of thymine hydroperoxides and are found as stable residues attached to the backbone of irradiated DNA (1). Urea residues are also in vitro replication blocks to DNA synthesis, however, since urea residues are totally noninstructive, synthesis is arrested one base before the damage (6,8). Apurinic/apyrimidinic (AP) sites are common DNA lesions produced by radiation and chemicals and are also formed as intermediates in repair processes initiated by DNA glycosylases (11). AP sites are noninstructive lesions that are also blocks to DNA replication in vitro (12). Thymine glycols, urea residues and AP sites are easily quantitated in DNA by enzymatic and immunochemical procedures (13,14). Most importantly, thymine glycols can be selectively introduced into DNA by osmium tetroxide oxidation (15,16); urea residues can be formed by alkali hydrolysis of thymine glycol-containing DNA (17); and AP sites can be introduced by heat/acid treatment (18).


Phosphodiester Backbone Replicative Block Lesion Bypass Hydroxyl Radical Attack Thymine Glycol 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    E. Teoule, C. Bert and A. Bonicel, Thymine fragment damage retained in the DNA polynucleotide chain after gamma-irradiation in aerated solutions, Radiat Res. 72: 190 (1977)PubMedCrossRefGoogle Scholar
  2. 2.
    R. Rajagopalan, R. J. Melamede, M. F. Laspia, B. F. Erlanger and S. S. Wallace, Properties of antibodies to thymine glycol, a product of the radiolysis of DNA, Radiat Res. 97: 499 (1984)PubMedCrossRefGoogle Scholar
  3. 3.
    L. H. Breimer and T. Lindahl, Thymine lesions produced by ionizing radiation in double stranded DNA, Biochemistry 24: 4018 (1985)PubMedCrossRefGoogle Scholar
  4. 4.
    S. A. Leadon and P. C. Hanawalt, Monoclonal antibody to DNA containing thymine glycol, Mutat Res DNA Repair Reports 112: 191 (1983)CrossRefGoogle Scholar
  5. 5.
    R. Cathcart, E. Schwiers, R. L. Saul and B. N. Ames, Thymine glycol and thymidine glycol in human and rat urine: a possible assay for oxidative DNA damage, Proc Natl Acad Sci. USA 81: 5633 (1984)PubMedCrossRefGoogle Scholar
  6. 6.
    H. Ide, Y. W. Kow and S. S. Wallace, Thymine glycols and urea residues in M13 DNA constitute replicative blocks in vitro, Nucleic Acids Res. 13: 8035 (1985)PubMedCrossRefGoogle Scholar
  7. 7.
    P. Rouet and J. M. Essigmann, Possible role for thymine glycol in the selective inhibition of DNA synthesis on oxidized DNA templates, Cancer Res. 45: 6113 (1985)PubMedGoogle Scholar
  8. 8.
    R. C. Hayes and J. E. LeClerc, Sequence dependence for bypass of thymine glycols in DNA by DNA polymerase I, Nucleic Acids Res. 14: 1045 (1986)PubMedCrossRefGoogle Scholar
  9. 9.
    J. M. Clark and G. P. Beardsley, Thymine glycol lesions terminate chain elongation by DNA polymerase in in vitro, Nucleic Acids Res. 14: 737 (1986)PubMedCrossRefGoogle Scholar
  10. 10.
    J. M. Clark and G. P. Beardsley, Functional effects of cis-thymine glycol lesions on DNA synthesis in vitro, Biochemistry 26: 5398 (1987)PubMedCrossRefGoogle Scholar
  11. 11.
    L. A. Loeb and B. D. Preston, Mutagenesis by apurinic/ apyrimidinic sites, Ann Rev Genet. 20: 201 (1986)PubMedCrossRefGoogle Scholar
  12. 12.
    D. Sagher and B. Strauss, Insertion of nucleotides opposite apurinic/apyrimidinic sites in deoxyribonucleic acid during in vitro synthesis: Uniqueness of adenine nucleotides, Biochemistry 22: 4518.Google Scholar
  13. 13.
    S. S. Wallace, Detection and repair of DNA base damage produced by ionizing radiation, Environ Mutagen. 5: 769 (1983)PubMedCrossRefGoogle Scholar
  14. 14.
    S. S. Wallace, The biological consequences of oxidized DNA bases, Br J Cancer 55, suppl. VIII: 118 (1987)Google Scholar
  15. 15.
    M. Beer, S. Stern, D. Carmalt and K. H. Mohlhenrich, Determination of base sequence in nucleic acids with the electron microscope. V. The thymine specific reactions of osmium tetroxide with deoxyribonucleic acid and components, Biochemistry 5: 2283 (1966)PubMedCrossRefGoogle Scholar
  16. 16.
    K. Frenkel, M. S. Goldstein and G. W. Teebor, Identification of the cis-thymine glycol moiety in chemically oxidized and gamma-irradiated deoxyribonucleic acid by high pressure liquid chromatography analysis, Biochemistry 20: 7566 (1984)CrossRefGoogle Scholar
  17. 17.
    Y. W. Kow and S. S. Wallace, Exonuclease III recognizes urea residues in oxidized DNA, Proc Natl Acad Sci. USA 82: 8354 (1985)PubMedCrossRefGoogle Scholar
  18. 18.
    T. Lindahl and A. Andersson, Rate of chain breakage at apurinic sites in double-stranded deoxyribonucleic acid, Biochemistry 11: 3618 (1972)PubMedCrossRefGoogle Scholar
  19. 19.
    B. Demple and S. Linn, DNA N-glycosylases and UV repair, Nature 287: 203 (1980)PubMedCrossRefGoogle Scholar
  20. 20.
    H. L. Katcher and S. S. Wallace, Characterization of the Escherichia coli X-ray endonuclease, endonuclease III, Biochemistry 22: 4071 (1983)PubMedCrossRefGoogle Scholar
  21. 21.
    B. Weiss, Endonuclease II of Escherichia coli is exonuclease III, J Biol Chem. 251: 1896 (1976)PubMedGoogle Scholar
  22. 22.
    S. Ljungquist, T. Lindahl and P. Howard-Flanders, Methyl methane sulfonate-sensitive mutant of Escherichia coli deficient in an endonuclease specific for apurinic sites in deoxyribonucleic acid, J Bacteriol. 126: 646 (1976)PubMedGoogle Scholar
  23. 23.
    S. Ljungquist and T. Lindahl, Relationship between Escherichia coli endonucleases specific for apurinic sites in DNA and exonuclease III, Nucleic Acids Res. 4: 2871 (1977)PubMedCrossRefGoogle Scholar
  24. 24.
    C. Milcarek and B. Weiss, Mutants of Escherichia coli with altered deoxyribonucleases, I. Isolation and characterization of mutants for exonuclease III, J Mol Biol. 68: 303 (1972)PubMedCrossRefGoogle Scholar
  25. 25.
    D. M. Yajko and B. Weiss, Mutations simultaneously affecting enconuclease II and exonuclease III in Escherichia coli, Proc Natl Acad Sci. USA 72: 688 (1975)PubMedCrossRefGoogle Scholar
  26. 26.
    B. Demple and J. Halbrook, Inducible repair of oxidative DNA damage in Escherichia coli, Nature 304: 466 (1983)PubMedCrossRefGoogle Scholar
  27. 27.
    B. F. Cunningham and B. Weiss, Endonuclease III (nth) mutants of Escherichia coli, Proc Natl Acad Sci. USA 82: 474 (1985)PubMedCrossRefGoogle Scholar
  28. 28.
    R. P. Cunningham, S. M. Saporito, S. G. Spitzer and B. Weiss, Enconuclease IV (nfo) mutant of Escherichia coli, J Bacteriol. 168: 1120.Google Scholar
  29. 29.
    Y. W. Kow and S. S. Wallace, Mechanism of action of endonuclease III from Escherichia coli, Biochemistry 26: 8200 (1987)PubMedCrossRefGoogle Scholar
  30. 30.
    V. Bailly and W. G. Verly, Escherichia coli endonuclease III is not an endonuclease but a ß-elimination catalyst, J Biochem. 242: 565 (1987)Google Scholar
  31. 31.
    H. R. Warner, B. F. Demple, W. A. Deutsch, C. M. Kane and S. Linn, Apurinic/apyrimidinic endonucleases in repair of pyrimidine dimers and other lesions in DNA, Proc Natl Acad Sci. USA 77: 4602 (1980)PubMedCrossRefGoogle Scholar
  32. 32.
    W. D. Taylor and W. Ginoza, Correlation of X-ray inactivation and strand scission in the replicative form of 0X-174 bacteriophage DNA, Proc Natl Acad Sci. USA 58: 1753 (1967)PubMedCrossRefGoogle Scholar
  33. 33.
    G. P. van der Schans, J. F. Bleichrodt and J. Blok, Contributions of various types of damage to inactivation of biologically active double-stranded circular DNA by gamma-radiation, Int J Radiat Biol. 23: 133 (1973)CrossRefGoogle Scholar
  34. 34.
    E. Moran and S. S. Wallace, The role of specific DNA base damage in the X-ray-induced inactivation of bacteriophage PM2, Mutat Res. 146: 229 (1985)PubMedCrossRefGoogle Scholar
  35. 35.
    J. L. Swinehart and P. A. Cerutti, Gamma-ray-induced thymine damage in the DNA in coliphage 0174 and in E. coli., Int J Radiat Biol. 27: 83 (1975)CrossRefGoogle Scholar
  36. 36.
    L. H. Breimer and T. Lindahl, DNA glycosylase activities for thymine residues damaged by ring-saturation, fragmentation, or ring-contraction are functions of endonuclease III in Escherichia coli, J Biol Chem. 259: 5543 (1984)PubMedGoogle Scholar
  37. 37.
    R. B. Weiss and N. J. Duker, Photoalkylated DNA and ultraviolet-irradiated DNA are incised at cytosines by endonuclease III, Nucl Acids Res. 14: 6621 (1986)PubMedCrossRefGoogle Scholar
  38. 38.
    P. W. Doetsch, D. E. Helland and W. A. Haseltine, Mechanism of action of a mammalian DNA repair endonuclease, Biochemistry 25: 2212 (1986)PubMedCrossRefGoogle Scholar
  39. 39.
    G. C. Walker, Mutagenesis and inducible response to deoxyribonucleic acid damage in Escherichia coli, Microbiol Rev. 48: 60 (1984)PubMedGoogle Scholar
  40. 40.
    P. M. Achey and C. F. Wright, Inducible repair of thymine ring saturation damage in ¢X174 DNA, Radiat Res. 93: 609 (1983)PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1989

Authors and Affiliations

  • Michael F. Laspia
    • 1
  • Susan S. Wallace
    • 1
  1. 1.Department of Microbiology and ImmunologyNew York Medical CollegeValhallaUSA

Personalised recommendations