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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1984 Dec;81(23):7397–7401. doi: 10.1073/pnas.81.23.7397

Escherichia coli DNA photolyase stimulates uvrABC excision nuclease in vitro.

A Sancar, K A Franklin, G B Sancar
PMCID: PMC392153  PMID: 6390436

Abstract

Pyrimidine dimers are the major photoproducts produced in cellular DNA upon UV irradiation. In Escherichia coli there are dark and photorepair mechanisms that eliminate the dimers from DNA and prevent their lethal and mutagenic effects. To determine whether these repair mechanisms act cooperatively or competitively in repairing DNA, we investigated the effects upon one another of DNA photolyase, which mediates photorepair, and uvrABC excision nuclease, an enzyme complex of the uvrABC gene products, which catalyzes nucleotide excision repair. We found that photolyase stimulates the removal of pyrimidine dimers but not other DNA adducts by uvrABC excision nuclease. The two subunits of uvrABC excision nuclease, the uvrA and uvrB proteins which together bind to the dimer region of DNA, had no effect on the activity of photolyase. T4 endonuclease V, which like photolyase is specific for pyrimidine dimers, was inhibited by photolyase, suggesting that these two proteins recognize the same or similar chemical structures in UV-irradiated DNA that are different from those recognized by uvrABC excision nuclease.

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Selected References

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  1. BOYCE R. P., HOWARD-FLANDERS P. RELEASE OF ULTRAVIOLET LIGHT-INDUCED THYMINE DIMERS FROM DNA IN E. COLI K-12. Proc Natl Acad Sci U S A. 1964 Feb;51:293–300. doi: 10.1073/pnas.51.2.293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. 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]
  4. Gordon L. K., Haseltine W. A. Comparison of the cleavage of pyrimidine dimers by the bacteriophage T4 and Micrococcus luteus UV-specific endonucleases. J Biol Chem. 1980 Dec 25;255(24):12047–12050. [PubMed] [Google Scholar]
  5. Harm H., Rupert C. S. Analysis of photoenzymatic repair of UV lesions in DNA by single light flashes. I. In vitro studies with Haemophilus influenzae transforming DNA and yeast photoreactivating enzyme. Mutat Res. 1968 Nov-Dec;6(3):355–370. doi: 10.1016/0027-5107(68)90053-5. [DOI] [PubMed] [Google Scholar]
  6. Jorns M. S., Sancar G. B., Sancar A. Identification of a neutral flavin radical and characterization of a second chromophore in Escherichia coli DNA photolyase. Biochemistry. 1984 Jun 5;23(12):2673–2679. doi: 10.1021/bi00307a021. [DOI] [PubMed] [Google Scholar]
  7. Kacinski B. M., Rupp W. D. E. coli uvrB protein binds to DNA in the presence of uvrA protein. Nature. 1981 Dec 3;294(5840):480–481. doi: 10.1038/294480a0. [DOI] [PubMed] [Google Scholar]
  8. Kacinski B. M., Sancar A., Rupp W. D. A general approach for purifying proteins encoded by cloned genes without using a functional assay: isolation of the uvrA gene product from radiolabeled maxicells. Nucleic Acids Res. 1981 Sep 25;9(18):4495–4508. doi: 10.1093/nar/9.18.4495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Nakabeppu Y., Sekiguchi M. Physical association of pyrimidine dimer DNA glycosylase and apurinic/apyrimidinic DNA endonuclease essential for repair of ultraviolet-damaged DNA. Proc Natl Acad Sci U S A. 1981 May;78(5):2742–2746. doi: 10.1073/pnas.78.5.2742. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Patrick M. H., Harm H. Substrate specificity of a bacterial UV endonuclease and the overlap with in vitro photoenzymatic repair. Photochem Photobiol. 1973 Nov;18(5):371–386. doi: 10.1111/j.1751-1097.1973.tb06437.x. [DOI] [PubMed] [Google Scholar]
  11. RUPERT C. S., GOODGAL S. H., HERRIOTT R. M. Photoreactivation in vitro of ultraviolet-inactivated Hemophilus influenzae transforming factor. J Gen Physiol. 1958 Jan 20;41(3):451–471. doi: 10.1085/jgp.41.3.451. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. 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]
  13. Rao S. N., Keepers J. W., Kollman P. The structure of d(CGCGAAT[]TCGCG) . d(CGCGAATTCGCG); the incorporation of a thymine photodimer into a B-DNA helix. Nucleic Acids Res. 1984 Jun 11;12(11):4789–4807. doi: 10.1093/nar/12.11.4789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ruffieux A., Schultz W. Dopaminergic activation of reticulata neurones in the substantia nigra. Nature. 1980 May 22;285(5762):240–241. doi: 10.1038/285240a0. [DOI] [PubMed] [Google Scholar]
  15. Rupp W. D., Wilde C. E., 3rd, Reno D. L., Howard-Flanders P. Exchanges between DNA strands in ultraviolet-irradiated Escherichia coli. J Mol Biol. 1971 Oct 14;61(1):25–44. doi: 10.1016/0022-2836(71)90204-x. [DOI] [PubMed] [Google Scholar]
  16. SETLOW R. B., CARRIER W. L. THE DISAPPEARANCE OF THYMINE DIMERS FROM DNA: AN ERROR-CORRECTING MECHANISM. Proc Natl Acad Sci U S A. 1964 Feb;51:226–231. doi: 10.1073/pnas.51.2.226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sancar A., Rupert C. S. Determination of plasmid molecular weights from ultraviolet sensitivities. Nature. 1978 Mar 30;272(5652):471–472. doi: 10.1038/272471a0. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Sancar A., Sancar G. B. Escherichia coli DNA photolyase is a flavoprotein. J Mol Biol. 1984 Jan 15;172(2):223–227. doi: 10.1016/s0022-2836(84)80040-6. [DOI] [PubMed] [Google Scholar]
  20. 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]
  21. Sancar G. B., Smith F. W., Sancar A. Identification and amplification of the E. coli phr gene product. Nucleic Acids Res. 1983 Oct 11;11(19):6667–6678. doi: 10.1093/nar/11.19.6667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Seeberg E., Steinum A. L. Purification and properties of the uvrA protein from Escherichia coli. Proc Natl Acad Sci U S A. 1982 Feb;79(4):988–992. doi: 10.1073/pnas.79.4.988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Setlow R. B., Carrier W. L. Pyrimidine dimers in ultraviolet-irradiated DNA's. J Mol Biol. 1966 May;17(1):237–254. doi: 10.1016/s0022-2836(66)80105-5. [DOI] [PubMed] [Google Scholar]
  25. WULFF D. L., RUPERT C. S. Disappearance of thymine photodimer in ultraviolet irradiated DNA upon treatment with a photoreactivating enzyme from baker's yeast. Biochem Biophys Res Commun. 1962 Apr 20;7:237–240. doi: 10.1016/0006-291x(62)90181-x. [DOI] [PubMed] [Google Scholar]
  26. Yamamoto K., Satake M., Shinagawa H. A multicopy phr-plasmid increases the ultraviolet resistance of a recA strain of Escherichia coli. Mutat Res. 1984 Jan;131(1):11–18. doi: 10.1016/0167-8817(84)90042-7. [DOI] [PubMed] [Google Scholar]
  27. 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]

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