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. 1992 Nov 15;89(22):11021–11025. doi: 10.1073/pnas.89.22.11021

Hydrolytic elimination of a mutagenic nucleotide, 8-oxodGTP, by human 18-kilodalton protein: sanitization of nucleotide pool.

J Y Mo 1, H Maki 1, M Sekiguchi 1
PMCID: PMC50475  PMID: 1332067

Abstract

8-Oxoguanine nucleotide can pair with cytosine and adenine nucleotides at almost equal efficiencies. Once 8-oxodGTP is formed in the cellular nucleotide pool, this mutagenic nucleotide is incorporated into DNA and would cause transversion mutations. The MutT protein of Escherichia coli possesses enzyme activity to hydrolyze 8-oxodGTP to the corresponding nucleoside monophosphate and thus may be responsible for preventing the occurrence of such mutations. Here we show that the human cell has an enzyme specifically hydrolyzing 8-oxodGTP in a fashion similar to that seen with MutT protein. The human 8-oxodGTPase has been found in cell-free extracts from Jurkat cells and purified > 400-fold. Analyses by gel filtration and gel electrophoresis revealed that the molecular mass of the native form of human 8-oxodGTPase is 18 kDa. Mg2+ ion is required for the enzyme action and the optimum pH for the reaction is pH 8.0. The enzyme hydrolyzes 8-oxodGTP to 8-oxodGMP with a Km value of 12.5 microM. dGTP and dATP are also degraded to dGMP and dAMP, respectively, with Km values 70 times greater than that for 8-oxodGTP. dTTP and dCTP are not hydrolyzed. These properties of the human 8-oxodGTPase are similar to those observed with the E. coli MutT protein, suggesting that the function of protecting the genetic information from the threat of endogenous oxygen radicals is widely distributed in organisms.

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Selected References

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

  1. Akiyama M., Maki H., Sekiguchi M., Horiuchi T. A specific role of MutT protein: to prevent dG.dA mispairing in DNA replication. Proc Natl Acad Sci U S A. 1989 Jun;86(11):3949–3952. doi: 10.1073/pnas.86.11.3949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ames B. N., Gold L. S. Endogenous mutagens and the causes of aging and cancer. Mutat Res. 1991 Sep-Oct;250(1-2):3–16. doi: 10.1016/0027-5107(91)90157-j. [DOI] [PubMed] [Google Scholar]
  3. Au K. G., Cabrera M., Miller J. H., Modrich P. Escherichia coli mutY gene product is required for specific A-G----C.G mismatch correction. Proc Natl Acad Sci U S A. 1988 Dec;85(23):9163–9166. doi: 10.1073/pnas.85.23.9163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bhatnagar S. K., Bessman M. J. Studies on the mutator gene, mutT of Escherichia coli. Molecular cloning of the gene, purification of the gene product, and identification of a novel nucleoside triphosphatase. J Biol Chem. 1988 Jun 25;263(18):8953–8957. [PubMed] [Google Scholar]
  5. Cabrera M., Nghiem Y., Miller J. H. mutM, a second mutator locus in Escherichia coli that generates G.C----T.A transversions. J Bacteriol. 1988 Nov;170(11):5405–5407. doi: 10.1128/jb.170.11.5405-5407.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cheng K. C., Cahill D. S., Kasai H., Nishimura S., Loeb L. A. 8-Hydroxyguanine, an abundant form of oxidative DNA damage, causes G----T and A----C substitutions. J Biol Chem. 1992 Jan 5;267(1):166–172. [PubMed] [Google Scholar]
  7. Duncan B. K., Weiss B. Specific mutator effects of ung (uracil-DNA glycosylase) mutations in Escherichia coli. J Bacteriol. 1982 Aug;151(2):750–755. doi: 10.1128/jb.151.2.750-755.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hennecke F., Kolmar H., Bründl K., Fritz H. J. The vsr gene product of E. coli K-12 is a strand- and sequence-specific DNA mismatch endonuclease. Nature. 1991 Oct 24;353(6346):776–778. doi: 10.1038/353776a0. [DOI] [PubMed] [Google Scholar]
  9. 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]
  10. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  11. Lu A. L., Chang D. Y. A novel nucleotide excision repair for the conversion of an A/G mismatch to C/G base pair in E. coli. Cell. 1988 Sep 9;54(6):805–812. doi: 10.1016/s0092-8674(88)91109-9. [DOI] [PubMed] [Google Scholar]
  12. Maki H., Sekiguchi M. MutT protein specifically hydrolyses a potent mutagenic substrate for DNA synthesis. Nature. 1992 Jan 16;355(6357):273–275. doi: 10.1038/355273a0. [DOI] [PubMed] [Google Scholar]
  13. Mo J. Y., Maki H., Sekiguchi M. Mutational specificity of the dnaE173 mutator associated with a defect in the catalytic subunit of DNA polymerase III of Escherichia coli. J Mol Biol. 1991 Dec 20;222(4):925–936. doi: 10.1016/0022-2836(91)90586-u. [DOI] [PubMed] [Google Scholar]
  14. Modrich P. DNA mismatch correction. Annu Rev Biochem. 1987;56:435–466. doi: 10.1146/annurev.bi.56.070187.002251. [DOI] [PubMed] [Google Scholar]
  15. Nghiem Y., Cabrera M., Cupples C. G., Miller J. H. The mutY gene: a mutator locus in Escherichia coli that generates G.C----T.A transversions. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2709–2713. doi: 10.1073/pnas.85.8.2709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Schaaper R. M., Danforth B. N., Glickman B. W. Mechanisms of spontaneous mutagenesis: an analysis of the spectrum of spontaneous mutation in the Escherichia coli lacI gene. J Mol Biol. 1986 May 20;189(2):273–284. doi: 10.1016/0022-2836(86)90509-7. [DOI] [PubMed] [Google Scholar]
  17. Shibutani S., Takeshita M., Grollman A. P. Insertion of specific bases during DNA synthesis past the oxidation-damaged base 8-oxodG. Nature. 1991 Jan 31;349(6308):431–434. doi: 10.1038/349431a0. [DOI] [PubMed] [Google Scholar]
  18. Shigenaga M. K., Gimeno C. J., Ames B. N. Urinary 8-hydroxy-2'-deoxyguanosine as a biological marker of in vivo oxidative DNA damage. Proc Natl Acad Sci U S A. 1989 Dec;86(24):9697–9701. doi: 10.1073/pnas.86.24.9697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Tchou J., Kasai H., Shibutani S., Chung M. H., Laval J., Grollman A. P., Nishimura S. 8-oxoguanine (8-hydroxyguanine) DNA glycosylase and its substrate specificity. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4690–4694. doi: 10.1073/pnas.88.11.4690. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Tkeshelashvili L. K., McBride T., Spence K., Loeb L. A. Mutation spectrum of copper-induced DNA damage. J Biol Chem. 1991 Apr 5;266(10):6401–6406. [PubMed] [Google Scholar]
  21. Wood R. D., Robins P., Lindahl T. Complementation of the xeroderma pigmentosum DNA repair defect in cell-free extracts. Cell. 1988 Apr 8;53(1):97–106. doi: 10.1016/0092-8674(88)90491-6. [DOI] [PubMed] [Google Scholar]
  22. Yamamoto F., Kasai H., Bessho T., Chung M. H., Inoue H., Ohtsuka E., Hori T., Nishimura S. Ubiquitous presence in mammalian cells of enzymatic activity specifically cleaving 8-hydroxyguanine-containing DNA. Jpn J Cancer Res. 1992 Apr;83(4):351–357. doi: 10.1111/j.1349-7006.1992.tb00114.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Yanofsky C., Cox E. C., Horn V. The unusual mutagenic specificity of an E. Coli mutator gene. Proc Natl Acad Sci U S A. 1966 Feb;55(2):274–281. doi: 10.1073/pnas.55.2.274. [DOI] [PMC free article] [PubMed] [Google Scholar]

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