Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1987 Feb 11;15(3):1227–1243. doi: 10.1093/nar/15.3.1227

Utilization of DNA photolyase, pyrimidine dimer endonucleases, and alkali hydrolysis in the analysis of aberrant ABC excinuclease incisions adjacent to UV-induced DNA photoproducts.

G M Myles, B Van Houten, A Sancar
PMCID: PMC340520  PMID: 3547334

Abstract

ABC excinuclease of Escherichia coli removes 6-4 photoproducts and pyrimidine dimers from DNA by making two single strand incisions, one 8 phosphodiester bonds 5' and another 4 or 5 phosphodiester bonds 3' to the lesion. We describe in this communication a method, which utilizes DNA photolyase from E. coli, pyrimidine dimer endonucleases from M. luteus and bacteriophage T4, and alkali hydrolysis, for analyzing the ABC excinuclease incision pattern corresponding to each of these photoproducts in a DNA fragment. On occasion, ABC excinuclease does not incise DNA exclusively 8 phosphodiester bonds 5' or 4 or 5 phosphodiester bonds 3' to the photoproduct. Both the nature of the adduct (6-4 photoproduct or pyrimidine dimer) and the sequence of neighboring nucleotides influence the incision pattern of ABC excinuclease. We show directly that photolyase stimulates the removal of pyrimidine dimers (but not 6-4 photoproducts) by the excinuclease. Also, photolyase does not repair CC pyrimidine dimers efficiently while it does repair TT or TC pyrimidine dimers.

Full text

PDF
1227

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arikan E., Kulkarni M. S., Thomas D. C., Sancar A. Sequences of the E. coli uvrB gene and protein. Nucleic Acids Res. 1986 Mar 25;14(6):2637–2650. doi: 10.1093/nar/14.6.2637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brash D. E., Franklin W. A., Sancar G. B., Sancar A., Haseltine W. A. Escherichia coli DNA photolyase reverses cyclobutane pyrimidine dimers but not pyrimidine-pyrimidone (6-4) photoproducts. J Biol Chem. 1985 Sep 25;260(21):11438–11441. [PubMed] [Google Scholar]
  3. Demple B., Linn S. DNA N-glycosylases and UV repair. Nature. 1980 Sep 18;287(5779):203–208. doi: 10.1038/287203a0. [DOI] [PubMed] [Google Scholar]
  4. Franklin W. A., Haseltine W. A. Removal of UV light-induced pyrimidine-pyrimidone(6-4) products from Escherichia coli DNA requires the uvrA, uvrB, and urvC gene products. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3821–3824. doi: 10.1073/pnas.81.12.3821. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Franklin W. A., Lo K. M., Haseltine W. A. Alkaline lability of fluorescent photoproducts produced in ultraviolet light-irradiated DNA. J Biol Chem. 1982 Nov 25;257(22):13535–13543. [PubMed] [Google Scholar]
  6. Haseltine W. A., Gordon L. K., Lindan C. P., Grafstrom R. H., Shaper N. L., Grossman L. Cleavage of pyrimidine dimers in specific DNA sequences by a pyrimidine dimer DNA-glycosylase of M. luteus. Nature. 1980 Jun 26;285(5767):634–641. doi: 10.1038/285634a0. [DOI] [PubMed] [Google Scholar]
  7. Husain I., Van Houten B., Thomas D. C., Sancar A. Sequences of Escherichia coli uvrA gene and protein reveal two potential ATP binding sites. J Biol Chem. 1986 Apr 15;261(11):4895–4901. [PubMed] [Google Scholar]
  8. Lippke J. A., Gordon L. K., Brash D. E., Haseltine W. A. Distribution of UV light-induced damage in a defined sequence of human DNA: detection of alkaline-sensitive lesions at pyrimidine nucleoside-cytidine sequences. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3388–3392. doi: 10.1073/pnas.78.6.3388. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  10. Radany E. H., Friedberg E. C. A pyrimidine dimer-DNA glycosylase activity associated with the v gene product of bacterophage T4. Nature. 1980 Jul 10;286(5769):182–185. doi: 10.1038/286182a0. [DOI] [PubMed] [Google Scholar]
  11. Sancar A., Franklin K. A., Sancar G. B. Escherichia coli DNA photolyase stimulates uvrABC excision nuclease in vitro. Proc Natl Acad Sci U S A. 1984 Dec;81(23):7397–7401. doi: 10.1073/pnas.81.23.7397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Sancar A., Franklin K. A., Sancar G., Tang M. S. Repair of psoralen and acetylaminofluorene DNA adducts by ABC excinuclease. J Mol Biol. 1985 Aug 20;184(4):725–734. doi: 10.1016/0022-2836(85)90316-x. [DOI] [PubMed] [Google Scholar]
  13. Sancar A., Rupp W. D. A novel repair enzyme: UVRABC excision nuclease of Escherichia coli cuts a DNA strand on both sides of the damaged region. Cell. 1983 May;33(1):249–260. doi: 10.1016/0092-8674(83)90354-9. [DOI] [PubMed] [Google Scholar]
  14. Sancar A., Smith F. W., Sancar G. B. Purification of Escherichia coli DNA photolyase. J Biol Chem. 1984 May 10;259(9):6028–6032. [PubMed] [Google Scholar]
  15. Sancar G. B., Sancar A., Rupp W. D. Sequences of the E. coli uvrC gene and protein. Nucleic Acids Res. 1984 Jun 11;12(11):4593–4608. doi: 10.1093/nar/12.11.4593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Seawell P. C., Smith C. A., Ganesan A. K. den V gene of bacteriophage T4 determines a DNA glycosylase specific for pyrimidine dimers in DNA. J Virol. 1980 Sep;35(3):790–796. doi: 10.1128/jvi.35.3.790-796.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Thomas D. C., Levy M., Sancar A. Amplification and purification of UvrA, UvrB, and UvrC proteins of Escherichia coli. J Biol Chem. 1985 Aug 15;260(17):9875–9883. [PubMed] [Google Scholar]
  18. Yeung A. T., Mattes W. B., Oh E. Y., Grossman L. Enzymatic properties of purified Escherichia coli uvrABC proteins. Proc Natl Acad Sci U S A. 1983 Oct;80(20):6157–6161. doi: 10.1073/pnas.80.20.6157. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES