Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

A specific partner for abasic damage in DNA

Nature volume 399, pages 704–708 (1999)Cite this article

Abstract

In most models of DNA replication, Watson–Crick hydrogen bonding drives the incorporation of nucleotides into the new strand of DNA and maintains the complementarity of bases with the template strand. Studies with nonpolar analogues of thymine and adenine, however, have shown that replication is still efficient in the absence of hydrogen bonds1,2,3,4. The replication of base pairs might also be influenced by steric exclusion, whereby inserted nucleotides need to be the correct size and shape to fit the active site against a template base5,6. A simple steric-exclusion model may not require Watson–Crick hydrogen bonding to explain the fidelity of replication, nor should canonical purine and pyrimidine shapes be necessary for enzymatic synthesis of a base pair if each can fit into the DNA double helix without steric strain6. Here we test this idea by using a pyrene nucleoside triphosphate (dPTP) in which the fluorescent ‘base’ is nearly as large as an entire Watson–Crick base pair. We show that the non-hydrogen-bonding dPTP is efficiently and specifically inserted by DNA polymerases opposite sites that lack DNA bases. The efficiency of this process approaches that of a natural base pair and the specificity is 102–104-fold. We use these properties to sequence abasic lesions in DNA, which are a common form of DNA damage in vivo7. In addition to their application in identifying such genetic lesions, our results show that neither hydrogen bonds nor purine and pyrimidine structures are required to form a base pair with high efficiency and selectivity. These findings confirm that steric complementarity is an important factor in the fidelity of DNA synthesis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Structures studied.
Figure 2: Polymerase insertion of dPTP into templates containing abasic sites.
Figure 3: Comparison of the steady-state efficiencies of dPTP insertion.
Figure 4: T7 DNA polymerase (exo-)-mediated sequencing of a template from the p53 gene sequence containing single and multiple abasic sites.

Similar content being viewed by others

References

  1. Moran, S., Ren, R. X.-F. & Kool, E. T. Athymidine triphosphate shape mimic lacking Watson–Crick pairing ability is replicated with high specificity. Proc. Natl Acad. Sci. USA 94, 10506–10511 (1997).

    Article  ADS  CAS  Google Scholar 

  2. Goodman, M. F. Hydrogen bonding revisited: geometric selection as a principal determinant of DNA replication fidelity. Proc. Natl Acad. Sci. USA 94, 10493–10495 (1997).

    Article  ADS  CAS  Google Scholar 

  3. Morales, J. C. & Kool, E. T. Efficient replication of a DNA base pair between non-hydrogen-bonded nucleoside analogues. Nature Struct. Biol. 5, 950–954 (1998).

    Article  CAS  Google Scholar 

  4. Moran, S., Ren, R. X.-F., Rumney, S. & Kool, E. T. Difluorotoluene, a nonpolar isostere of thymine, codes specifically and efficiently for adenine in DNA replication. J. Am. Chem. Soc. 119, 2056–2057 (1997).

    Article  CAS  Google Scholar 

  5. Goodman, M. F., Creighton, S., Bloom, L. B. & Petruska, J. Biochemical basis of DNA replication fidelity. Crit. Rev. Biochem. Mol. Biol. 28, 83–126 (1993).

    Article  CAS  Google Scholar 

  6. Kool, E. T. Replication of non-hydrogen bonded bases by DNA polymerases: a mechanism for steric matching. Biopolymers (Nucleic Acid Sciences) 48, 3–17 (1998).

    Article  CAS  Google Scholar 

  7. Loeb, L. A. & Preston, B. D. Mutagenesis by apurinic/apyrimidinic sites. Annu. Rev. Genet. 20, 201–230 (1986).

    Article  CAS  Google Scholar 

  8. Ren, R. X.-F., Chaudhuri, N. C., Paris, P. L., Rumney, S. & Kool, E. T. Naphthalene, phenanthrene, and pyrene as DNA base analogues: synthesis, structure, and fluorescence in DNA. J. Am. Chem. Soc. 118, 7671–7678 (1996).

    Article  CAS  Google Scholar 

  9. Matray, T. J. & Kool, E. T. Selective and stable DNA base pairing without hydrogen bonds. J. Am. Chem. Soc. 120, 6191–6192 (1998).

    Article  CAS  Google Scholar 

  10. Mishra, M. C. & Broom, A. D. Anovel synthesis of nucleoside 5′ triphosphates. J. Chem. Soc., Chem. Commun. 1276–1277 (1991).

  11. Hoard, D. E. & Ott, D. G. Conversion of mono- and oligodeoxyribonucleotides to 5′-triphosphates. J. Am. Chem. Soc. 87, 1785–1788 (1965).

    Article  CAS  Google Scholar 

  12. Millican, T. A. et al. . Synthesis and biophysical studies of short oligodeoxynucleotides with novel modifications. Nucleic Acids Res. 12, 7435–7453 (1984).

    Article  CAS  Google Scholar 

  13. Takeshita, M., Chang,,, C.,, Johnson, F., Will, S. & Grollman, A. P. Oligodeoxy-nucleotides containing synthetic abasic sites. Model substrates for DNA polymerases and apurinic/apyrimidinic endonucleases. J. Biol. Chem. 262, 10171–10179 (1987).

    CAS  PubMed  Google Scholar 

  14. Randall, S. K., Eritja, R., Kaplan, B. E., Petruska, J. & Goodman, M. F. Nucleotide insertion kinetics opposite abasic lesions in DNA. J. Biol. Chem. 262, 6864–6870 (1987).

    CAS  PubMed  Google Scholar 

  15. Sagher, D. & Strauss, B. Insertion of nucleotides opposite apurinic/apyrimidinic sites in deoxyribonucleic acid during in vitro synthesis: uniqueness of adenine nucleotides. Biochemistry 22, 4518–4526 (1983).

    Article  CAS  Google Scholar 

  16. Schaaper, R. M., Kunkel, T. A. & Loeb, L. A. Infidelity of DNA synthesis associated with bypass of apurinic sites. Proc. Natl Acad. Sci. USA 80, 487–491 (1983).

    Article  ADS  CAS  Google Scholar 

  17. Lawrence, C. W., Borden, A., Banerjee, S. K. & LeClerc, J. E. Mutation frequency and spectrum resulting from a single abasic site in a single-stranded vector. Nucleic Acids Res. 18, 2153–2157 (1990).

    Article  CAS  Google Scholar 

  18. Paz-Elizur, T., Takeshita, M. & Livneh, Z. Mechanism of bypass synthesis through an abasic site analog by DNA polymerase I. Biochemistry 36, 1766–1773 (1997).

    Article  CAS  Google Scholar 

  19. Doublié, S., Tabor, S., Long, A. M., Richardson, C. C. & Ellenberger, T. Crystal structure of a bacteriophage T7 DNA replication complex at 2.2 å resolution. Nature 391, 251–258 (1998).

    Article  ADS  Google Scholar 

  20. Kiefer, J. R., Mao, C., Braman, J. C. & Beese, L. S. Visualizing DNA replication in a catalytically active Bacillus DNA polymerase crystal. Nature 391, 304–307 (1998).

    Article  ADS  CAS  Google Scholar 

  21. Huang, H., Chopra, R., Verdine, G. L. & Harrison, S. C. Structure of a covalently trapped catalytic complex of HIV-1 reverse transcriptase: implications for drug resistance. Science 282, 1669–1675 (1998).

    Article  ADS  CAS  Google Scholar 

  22. Diederichsen, U. Selectivity of DNA replication: the importance of geometry over hydrogen bonding. Angew. Chem. 37, 1655–1657 (1998).

    Article  CAS  Google Scholar 

  23. Fygenson, D. K. & Goodman, M. F. Appendix. Gel kinetic analysis of polymerase fidelity in the presence of multiple enzyme DNA encounters. J. Biol. Chem. 272, 27931–27935 (1997).

    Article  CAS  Google Scholar 

  24. Shibutani, S., Takeshita, M. & Grollman, A. P. Translesional synthesis on DNA templates containing a single abasic site. A mechanistic study of the “A rule”. J. Biol. Chem. 272, 13916–13921 (1997).

    Article  CAS  Google Scholar 

  25. Lindahl, T. & Nyberg, B. Rate of depurination of native deoxyribonucleic acid. Biochemistry 11, 3610–3618 (1972).

    Article  CAS  Google Scholar 

  26. Weiss, B. & Grossman, L. Phosphodiesterases involved in DNA repair. Adv. Enzymol. Rel. Areas Mol. Biol. 60, 1–34 (1972).

    Google Scholar 

  27. Ide, H. et al. . Synthesis and damage specificity of a novel probe for the detection of abasic sites in DNA. Biochemistry 32, 8276–8283 (1993).

    Article  CAS  Google Scholar 

  28. Maulik, G. et al. . Novel non-isotopic detection of MutY enzyme-recognized mismatches in DNA via ultrasensitive detection of aldehydes. Nucleic Acids Res. 27, 1316–1322 (1999).

    Article  CAS  Google Scholar 

  29. Denissenko, M. F., Pao, A., Tang, M. & Pfeifer, G. P. Preferential formation of benzo[a ]pyrene adducts at lung cancer mutational hotspots in p53. Science 274, 430–432 (1996).

    Article  ADS  CAS  Google Scholar 

  30. Paris, P. L., Langenhan, J. & Kool, E. T. Probing DNA sequences in solution with a monomer–excimer fluorescence color change. Nucleic Acids Res. 26, 3789–3793 (1998).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the NIH and from the US Army Research Office.

Author information

Authors and Affiliations

  1. Department of Chemistry, University of Rochester, Rochester, 14627, New York, USA

    Tracy J. Matray & Eric T. Kool

Authors
  1. Tracy J. Matray
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Eric T. Kool
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Eric T. Kool.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Matray, T., Kool, E. A specific partner for abasic damage in DNA. Nature 399, 704–708 (1999). https://doi.org/10.1038/21453

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/21453

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing