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. 1998 Mar 15;330(Pt 3):1059–1067. doi: 10.1042/bj3301059

The protein oxidation product 3,4-dihydroxyphenylalanine (DOPA) mediates oxidative DNA damage.

B Morin 1, M J Davies 1, R T Dean 1
PMCID: PMC1219245  PMID: 9494069

Abstract

A major product of hydroxy-radical addition to tyrosine is 3, 4-dihydroxyphenylalanine (DOPA) which has reducing properties. Protein-bound DOPA (PB-DOPA) has been shown to be a major component of the stable reducing species formed during protein oxidation under several conditions. The aim of the present work was to investigate whether DOPA, and especially PB-DOPA, can mediate oxidative damage to DNA. We chose to generate PB-DOPA using mushroom tyrosinase, which catalyses the hydroxylation of tyrosine residues in protein. This permitted us to study the reactions of PB-DOPA in the virtual absence of other protein-bound oxidation products. The formation of two oxidation products of DNA, 8-oxo-7,8-dihydro-2'-deoxyguanosine (8oxodG) and 5-hydroxy-2'-deoxycytidine (5OHdC), were studied with a novel HPLC using gradient elution and an electrochemical detection method, which allowed the detection of both DNA modifications in a single experiment. We found that exposure of calf thymus DNA to DOPA or PB-DOPA resulted in the formation of 8oxodG and 5OHdC, with the former predominating. The formation of these DNA oxidation products by either DOPA or PB-DOPA depended on the presence of oxygen, and also on the presence and on the concentration of transition metal ions, with copper being more effective than iron. The yields of 8oxodG and 5OHdC increased with DOPA concentration in proteins. Thus PB-DOPA was able to promote further radical-generating events, which then transferred damage to other biomolecules such as DNA.

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Selected References

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  1. Aruoma O. I., Halliwell B., Gajewski E., Dizdaroglu M. Copper-ion-dependent damage to the bases in DNA in the presence of hydrogen peroxide. Biochem J. 1991 Feb 1;273(Pt 3):601–604. doi: 10.1042/bj2730601. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bryan S. E., Vizard D. L., Beary D. A., LaBiche R. A., Hardy K. J. Partitioning of zinc and copper within subnuclear nucleoprotein particles. Nucleic Acids Res. 1981 Nov 11;9(21):5811–5823. doi: 10.1093/nar/9.21.5811. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chevion M. A site-specific mechanism for free radical induced biological damage: the essential role of redox-active transition metals. Free Radic Biol Med. 1988;5(1):27–37. doi: 10.1016/0891-5849(88)90059-7. [DOI] [PubMed] [Google Scholar]
  4. Chiu S. M., Xue L. Y., Friedman L. R., Oleinick N. L. Differential dependence on chromatin structure for copper and iron ion induction of DNA double-strand breaks. Biochemistry. 1995 Feb 28;34(8):2653–2661. doi: 10.1021/bi00008a032. [DOI] [PubMed] [Google Scholar]
  5. Davies M. J., Fu S., Dean R. T. Protein hydroperoxides can give rise to reactive free radicals. Biochem J. 1995 Jan 15;305(Pt 2):643–649. doi: 10.1042/bj3050643. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Davies M. J. Protein and peptide alkoxyl radicals can give rise to C-terminal decarboxylation and backbone cleavage. Arch Biochem Biophys. 1996 Dec 1;336(1):163–172. doi: 10.1006/abbi.1996.0545. [DOI] [PubMed] [Google Scholar]
  7. Dean R. T., Fu S., Stocker R., Davies M. J. Biochemistry and pathology of radical-mediated protein oxidation. Biochem J. 1997 May 15;324(Pt 1):1–18. doi: 10.1042/bj3240001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dean R. T., Gebicki J., Gieseg S., Grant A. J., Simpson J. A. Hypothesis: a damaging role in aging for reactive protein oxidation products? Mutat Res. 1992 Sep;275(3-6):387–393. doi: 10.1016/0921-8734(92)90041-m. [DOI] [PubMed] [Google Scholar]
  9. Dean R. T., Gieseg S., Davies M. J. Reactive species and their accumulation on radical-damaged proteins. Trends Biochem Sci. 1993 Nov;18(11):437–441. doi: 10.1016/0968-0004(93)90145-d. [DOI] [PubMed] [Google Scholar]
  10. Devasagayam T. P., Steenken S., Obendorf M. S., Schulz W. A., Sies H. Formation of 8-hydroxy(deoxy)guanosine and generation of strand breaks at guanine residues in DNA by singlet oxygen. Biochemistry. 1991 Jun 25;30(25):6283–6289. doi: 10.1021/bi00239a029. [DOI] [PubMed] [Google Scholar]
  11. Douki T., Delatour T., Paganon F., Cadet J. Measurement of oxidative damage at pyrimidine bases in gamma-irradiated DNA. Chem Res Toxicol. 1996 Oct-Nov;9(7):1145–1151. doi: 10.1021/tx960095b. [DOI] [PubMed] [Google Scholar]
  12. Fazal F., Rahman A., Greensill J., Ainley K., Hadi S. M., Parish J. H. Strand scission in DNA by quercetin and Cu(II): identification of free radical intermediates and biological consequences of scission. Carcinogenesis. 1990 Nov;11(11):2005–2008. doi: 10.1093/carcin/11.11.2005. [DOI] [PubMed] [Google Scholar]
  13. Floyd R. A., Watson J. J., Wong P. K., Altmiller D. H., Rickard R. C. Hydroxyl free radical adduct of deoxyguanosine: sensitive detection and mechanisms of formation. Free Radic Res Commun. 1986;1(3):163–172. doi: 10.3109/10715768609083148. [DOI] [PubMed] [Google Scholar]
  14. Floyd R. A., West M. S., Eneff K. L., Hogsett W. E., Tingey D. T. Hydroxyl free radical mediated formation of 8-hydroxyguanine in isolated DNA. Arch Biochem Biophys. 1988 Apr;262(1):266–272. doi: 10.1016/0003-9861(88)90188-9. [DOI] [PubMed] [Google Scholar]
  15. Fu S. L., Dean R. T. Structural characterization of the products of hydroxyl-radical damage to leucine and their detection on proteins. Biochem J. 1997 May 15;324(Pt 1):41–48. doi: 10.1042/bj3240041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fu S., Gebicki S., Jessup W., Gebicki J. M., Dean R. T. Biological fate of amino acid, peptide and protein hydroperoxides. Biochem J. 1995 Nov 1;311(Pt 3):821–827. doi: 10.1042/bj3110821. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fu S., Hick L. A., Sheil M. M., Dean R. T. Structural identification of valine hydroperoxides and hydroxides on radical-damaged amino acid, peptide, and protein molecules. Free Radic Biol Med. 1995 Sep;19(3):281–292. doi: 10.1016/0891-5849(95)00021-o. [DOI] [PubMed] [Google Scholar]
  18. Gebicki S., Gebicki J. M. Formation of peroxides in amino acids and proteins exposed to oxygen free radicals. Biochem J. 1993 Feb 1;289(Pt 3):743–749. doi: 10.1042/bj2890743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Geierstanger B. H., Kagawa T. F., Chen S. L., Quigley G. J., Ho P. S. Base-specific binding of copper(II) to Z-DNA. The 1.3-A single crystal structure of d(m5CGUAm5CG) in the presence of CuCl2. J Biol Chem. 1991 Oct 25;266(30):20185–20191. doi: 10.2210/pdb1d40/pdb. [DOI] [PubMed] [Google Scholar]
  20. George A. M., Sabovljev S. A., Hart L. E., Cramp W. A., Harris G., Hornsey S. DNA quaternary structure in the radiation sensitivity of human lymphocytes--a proposed role of copper. Br J Cancer Suppl. 1987 Jun;8:141–144. [PMC free article] [PubMed] [Google Scholar]
  21. Gieseg S. P., Simpson J. A., Charlton T. S., Duncan M. W., Dean R. T. Protein-bound 3,4-dihydroxyphenylalanine is a major reductant formed during hydroxyl radical damage to proteins. Biochemistry. 1993 May 11;32(18):4780–4786. doi: 10.1021/bi00069a012. [DOI] [PubMed] [Google Scholar]
  22. Halliwell B., Gutteridge J. M. Formation of thiobarbituric-acid-reactive substance from deoxyribose in the presence of iron salts: the role of superoxide and hydroxyl radicals. FEBS Lett. 1981 Jun 15;128(2):347–352. doi: 10.1016/0014-5793(81)80114-7. [DOI] [PubMed] [Google Scholar]
  23. Halliwell B., Gutteridge J. M. Oxygen toxicity, oxygen radicals, transition metals and disease. Biochem J. 1984 Apr 1;219(1):1–14. doi: 10.1042/bj2190001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hearing V. J., Jr, Ekel T. M., Montague P. M., Nicholson J. M. Mammalin tyrosinase. Stoichiometry and measurement of reaction products. Biochim Biophys Acta. 1980 Feb 14;611(2):251–268. doi: 10.1016/0005-2744(80)90061-3. [DOI] [PubMed] [Google Scholar]
  25. Hicks M., Delbridge L., Yue D. K., Reeve T. S. Catalysis of lipid peroxidation by glucose and glycosylated collagen. Biochem Biophys Res Commun. 1988 Mar 15;151(2):649–655. doi: 10.1016/s0006-291x(88)80330-9. [DOI] [PubMed] [Google Scholar]
  26. Husain S., Hadi S. M. Strand scission in DNA induced by L-DOPA in the presence of Cu(II). FEBS Lett. 1995 May 1;364(1):75–78. doi: 10.1016/0014-5793(95)00365-g. [DOI] [PubMed] [Google Scholar]
  27. Ito S., Kato T., Shinpo K., Fujita K. Oxidation of tyrosine residues in proteins by tyrosinase. Formation of protein-bonded 3,4-dihydroxyphenylalanine and 5-S-cysteinyl-3,4-dihydroxyphenylalanine. Biochem J. 1984 Sep 1;222(2):407–411. doi: 10.1042/bj2220407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kasai H., Crain P. F., Kuchino Y., Nishimura S., Ootsuyama A., Tanooka H. Formation of 8-hydroxyguanine moiety in cellular DNA by agents producing oxygen radicals and evidence for its repair. Carcinogenesis. 1986 Nov;7(11):1849–1851. doi: 10.1093/carcin/7.11.1849. [DOI] [PubMed] [Google Scholar]
  29. Kasai H., Nishimura S. Hydroxylation of deoxyguanosine at the C-8 position by ascorbic acid and other reducing agents. Nucleic Acids Res. 1984 Feb 24;12(4):2137–2145. doi: 10.1093/nar/12.4.2137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kelman D. J., Mason R. P. The myoglobin-derived radical formed on reaction of metmyoglobin with hydrogen peroxide is not a tyrosine peroxyl radical. Free Radic Res Commun. 1992;16(1):27–33. doi: 10.3109/10715769209049156. [DOI] [PubMed] [Google Scholar]
  31. Lewis C. D., Laemmli U. K. Higher order metaphase chromosome structure: evidence for metalloprotein interactions. Cell. 1982 May;29(1):171–181. doi: 10.1016/0092-8674(82)90101-5. [DOI] [PubMed] [Google Scholar]
  32. Løvstad R. A. Copper catalyzed oxidation of ascorbate (vitamin C). Inhibitory effect of catalase, superoxide dismutase, serum proteins (ceruloplasmin, albumin, apotransferrin) and amino acids. Int J Biochem. 1987;19(4):309–313. doi: 10.1016/0020-711x(87)90003-6. [DOI] [PubMed] [Google Scholar]
  33. Malkin R., Malmström B. G., Vänngård T. The reversible removal of one specific copper(II) from fungal laccase. Eur J Biochem. 1969 Jan;7(2):253–259. doi: 10.1111/j.1432-1033.1969.tb19600.x. [DOI] [PubMed] [Google Scholar]
  34. Muir Wood P. The redox potential of the system oxygen--superoxide. FEBS Lett. 1974 Aug 15;44(1):22–24. doi: 10.1016/0014-5793(74)80297-8. [DOI] [PubMed] [Google Scholar]
  35. Pacifici R. E., Davies K. J. Protein, lipid and DNA repair systems in oxidative stress: the free-radical theory of aging revisited. Gerontology. 1991;37(1-3):166–180. doi: 10.1159/000213257. [DOI] [PubMed] [Google Scholar]
  36. Park J. W., Floyd R. A. Lipid peroxidation products mediate the formation of 8-hydroxydeoxyguanosine in DNA. Free Radic Biol Med. 1992;12(4):245–250. doi: 10.1016/0891-5849(92)90111-s. [DOI] [PubMed] [Google Scholar]
  37. Prota G. Recent advances in the chemistry of melanogenesis in mammals. J Invest Dermatol. 1980 Jul;75(1):122–127. doi: 10.1111/1523-1747.ep12521344. [DOI] [PubMed] [Google Scholar]
  38. Rahman A., Shahabuddin, Hadi S. M., Parish J. H., Ainley K. Strand scission in DNA induced by quercetin and Cu(II): role of Cu(I) and oxygen free radicals. Carcinogenesis. 1989 Oct;10(10):1833–1839. doi: 10.1093/carcin/10.10.1833. [DOI] [PubMed] [Google Scholar]
  39. Schneider J. E., Price S., Maidt L., Gutteridge J. M., Floyd R. A. Methylene blue plus light mediates 8-hydroxy 2'-deoxyguanosine formation in DNA preferentially over strand breakage. Nucleic Acids Res. 1990 Feb 11;18(3):631–635. doi: 10.1093/nar/18.3.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Simpson J. A., Dean R. T. Stimulatory and inhibitory actions of proteins and amino acids on copper-catalysed free radical generation in the bulk phase. Free Radic Res Commun. 1990;10(4-5):303–312. doi: 10.3109/10715769009149899. [DOI] [PubMed] [Google Scholar]
  41. Simpson J. A., Gieseg S. P., Dean R. T. Free radical and enzymatic mechanisms for the generation of protein bound reducing moieties. Biochim Biophys Acta. 1993 Feb 13;1156(2):190–196. doi: 10.1016/0304-4165(93)90135-u. [DOI] [PubMed] [Google Scholar]
  42. Simpson J. A., Narita S., Gieseg S., Gebicki S., Gebicki J. M., Dean R. T. Long-lived reactive species on free-radical-damaged proteins. Biochem J. 1992 Mar 15;282(Pt 3):621–624. doi: 10.1042/bj2820621. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Soszynski M., Filipiak A., Bartosz G., Gebicki J. M. Effect of amino acid peroxides on the erythrocyte. Free Radic Biol Med. 1996;20(1):45–51. doi: 10.1016/0891-5849(95)02015-2. [DOI] [PubMed] [Google Scholar]
  44. Soszyński M., Skalski Z., Pułaski L., Bartosz G. Peroxides inhibit the glutathione S-conjugate pump. Biochem Mol Biol Int. 1995 Oct;37(3):537–545. [PubMed] [Google Scholar]
  45. Spencer J. P., Jenner A., Aruoma O. I., Evans P. J., Kaur H., Dexter D. T., Jenner P., Lees A. J., Marsden D. C., Halliwell B. Intense oxidative DNA damage promoted by L-dopa and its metabolites. Implications for neurodegenerative disease. FEBS Lett. 1994 Oct 24;353(3):246–250. doi: 10.1016/0014-5793(94)01056-0. [DOI] [PubMed] [Google Scholar]
  46. Stadtman E. R., Berlett B. S. Fenton chemistry. Amino acid oxidation. J Biol Chem. 1991 Sep 15;266(26):17201–17211. [PubMed] [Google Scholar]
  47. WEIL-MALHERBE H., BONE A. D. The chemical estimation of adrenaline-like substances in blood. Biochem J. 1952 Jun;51(3):311–318. doi: 10.1042/bj0510311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wagner J. R., Hu C. C., Ames B. N. Endogenous oxidative damage of deoxycytidine in DNA. Proc Natl Acad Sci U S A. 1992 Apr 15;89(8):3380–3384. doi: 10.1073/pnas.89.8.3380. [DOI] [PMC free article] [PubMed] [Google Scholar]

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