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Catalytically Competent Conformation of the Active Site of Human 8-Oxoguanine-DNA Glycosylase

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Abstract

8-Oxoguanine-DNA N-glycosylase (OGG1) is a eukaryotic DNA repair enzyme responsible for the removal of 8-oxoguanine (oxoG), one of the most abundant oxidative DNA lesions. OGG1 catalyzes two successive reactions - N-gly-cosidic bond hydrolysis (glycosylase activity) and DNA strand cleavage on the 3’-side of the lesion by ß-elimination (lyase activity). The enzyme also exhibits lyase activity with substrates containing apurinic/apyrimidinic (AP) sites (deoxyribose moieties lacking the nucleobase). OGG1 is highly specific for the base opposite the lesion, efficiently excising oxoG and cleaving AP sites located opposite to C, but not opposite to A. The activity is also profoundly decreased by amino acid changes that sterically interfere with oxoG binding in the active site of the enzyme after the lesion is everted from the DNA duplex. Earlier, the molecular dynamics approach was used to study the conformational dynamics of such human OGG1 mutants in complexes with the oxoG:C-containing substrate DNA, and the population density of certain conformers of two OGG1 catalytic residues, Lys249 and Asp268, was suggested to determine the enzyme activity. Here, we report the study of molecular dynamics of human OGG1 bound to the oxoG:A-containing DNA and OGG1 mutants bound to the AP:C-con-taining DNA. We showed that the enzyme low activity is associated with a decrease in the populations of Lys249 and Asp268 properly configured for catalysis. The experimentally measured rate constants for the OGG1 mutants show a good agreement with the models. We conclude that the enzymatic activity of OGG1 is determined majorly by the population density of the catalytically competent conformations of the active site residues Lys249 and Asp268.

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Abbreviations

AP site:

apurinic/apyrimidinic site

MD:

molecular dynamics

OGG1:

8-oxoguanine-DNA N-glycosylase

oxodG:

8-oxo-2′-deoxyguanosine

oxoG:

8-oxoguanine

WT:

wild type

References

  1. Von Sonntag, C. (2006) Free-Radical-Induced DNA Damage and Its Repair: A Chemical Perspective, Springer, Berlin-Heidelberg.

    Book  Google Scholar 

  2. Halliwell, B., and Gutteridge, J. M. C. (2007) Free Radicals in Biology and Medicine, 4th Edn., Oxford University Press, Oxford.

    Google Scholar 

  3. Friedberg, E. C., Walker, G. C., Siede, W., Wood, R. D., Schultz, R. A., and Ellenberger, T. (2006) DNA Repair and Mutagenesis, ASM Press, Washington, D. C.

    Google Scholar 

  4. Culp, S. J., Cho, B. P., Kadlubar, F. F., and Evans, F. E. (1989) Structural and conformational analyses of 8-hydroxy-2’-deoxyguanosine, Chem. Res. Toxicol., 2, 416–422, doi: 10.1021/tx00012a010.

    Article  CAS  PubMed  Google Scholar 

  5. Kouchakdjian, M., Bodepudi, V., Shibutani, S., Eisenberg, M., Johnson, F., Grollman, A. P., and Patel, D. J. (1991) NMR structural studies of the ionizing radiation adduct 7-hydro-8-oxodeoxyguanosine (8-oxo-7H-dG) opposite deoxyadenosine in a DNA duplex. 8-Oxo-7H-dG(syn)• dA(anti) alignment at lesion site, Biochemistry, 30, 1403–1412, doi: 10.1021/bi00219a034.

    Article  CAS  PubMed  Google Scholar 

  6. McAuley-Hecht, K. E., Leonard, G. A., Gibson, N. J., Thomson, J. B., Watson, W. P., Hunter, W. N., and Brown, T. (1994) Crystal structure of a DNA duplex containing 8-hydroxydeoxyguanine-adenine base pairs, Biochemistry, 33, 10266–10270, doi: 10.1021/bi00200a006.

    Article  CAS  PubMed  Google Scholar 

  7. Lipscomb, L. A., Peek, M. E., Morningstar, M. L., Verghis, S. M., Miller, E. M., Rich, A., Essigmann, J. M., and Williams, L. D. (1995) X-Ray structure of a DNA decamer containing 7,8-dihydro-8-oxoguanine, Proc. Natl. Acad. Sci. USA, 92, 719–723, doi: 10.1073/pnas.92.3.719.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Shibutani, S., Takeshita, M., and Grollman, A. P. (1991) Insertion of specific bases during DNA synthesis past the oxidation-damaged base 8-oxodG, Nature, 349, 431–434, doi: 10.1038/349431a0.

    Article  CAS  PubMed  Google Scholar 

  9. Grollman, A. P., and Moriya, M. (1993) Mutagenesis by 8-oxoguanine: an enemy within, Trends Genet., 9, 246–249, doi: 10.1016/0168-9525(93)90089-Z.

    Article  CAS  PubMed  Google Scholar 

  10. ESCODD (European Standards Committee on Oxidative DNA Damage), Gedik, C. M., and Collins, A. (2005) Establishing the background level of base oxidation in human lymphocyte DNA: results of an interlaboratory validation study, FASEB J., 19, 82–84, doi: 10.1096/fj.04-1767fje.

    Article  CAS  Google Scholar 

  11. Atamna, H., Cheung, I., and Ames, B. N. (2000) A method for detecting abasic sites in living cells: age-dependent changes in base excision repair, Proc. Natl. Acad. Sci. USA, 97, 686–691, doi: 10.1073/pnas.97.2.686.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Auffret van der Kemp, P., Thomas, D., Barbey, R., de Oliveira, R., and Boiteux, S. (1996) Cloning and expression in Escherichia coli of the OGG1 gene of Saccharomyces cere-visiae, which codes for a DNA glycosylase that excises 7,8-dihydro-8-oxoguanine and 2,6-diamino-4-hydroxy-5-N-methylformamidopyrimidine, Proc. Natl. Acad. Sci. USA, 93, 5197–5202, doi: 10.1073/pnas.93.11.5197.

    Article  Google Scholar 

  13. Rosenquist, T. A., Zharkov, D. O., and Grollman, A. P. (1997) Cloning and characterization of a mammalian 8-oxoguanine DNA glycosylase, Proc. Natl. Acad. Sci. USA, 94, 7429–7434, doi: 10.1073/pnas.94.14.7429.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Radicella, J. P., Dherin, C., Desmaze, C., Fox, M. S., and Boiteux, S. (1997) Cloning and characterization of hOGG1, a human homolog of the OGG1 gene of Saccharomyces cere-visiae, Proc. Natl. Acad. Sci. USA, 94, 8010–8015, doi: 10.1073/pnas.94.15.8010.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Roldan-Arjona, T., Wei, Y.-F., Carter, K. C., Klungland, A., Anselmino, C., Wang, R.-P., Augustus, M., and Lindahl, T. (1997) Molecular cloning and functional expression of a human cDNA encoding the antimutator enzyme 8-hydroxyguanine-DNA glycosylase, Proc. Natl. Acad. Sci. USA, 94, 8016–8020, doi: 10.1073/pnas. 94.15.8016.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Tchou, J., Kasai, H., Shibutani, S., Chung, M.-H., Laval, J., Grollman, A. P., and Nishimura, S. (1991) 8-Oxoguanine (8-hydroxyguanine) DNA glycosylase and its substrate specificity, Proc. Natl. Acad. Sci. USA, 88, 4690–4694, doi: 10.1073/pnas.88.11.4690.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Boiteux, S., Gajewski, E., Laval, J., and Dizdaroglu, M. (1992) Substrate specificity of the Escherichia coli Fpg protein (formamidopyrimidine-DNA glycosylase): excision of purine lesions in DNA produced by ionizing radiation or photosensitization, Biochemistry, 31, 106–110, doi: 10.1021/bi00116a016.

    Article  CAS  PubMed  Google Scholar 

  18. Nash, H. M., Lu, R., Lane, W. S., and Verdine, G. L. (1997) The critical active-site amine of the human 8-oxoguanine DNA glycosylase, hOgg1: direct identification, ablation and chemical reconstitution, Chem. Biol., 4, 693–702, doi: 10.1016/S1074-5521(97)90225-8.

    Article  CAS  PubMed  Google Scholar 

  19. Zharkov, D. O., Rosenquist, T. A., Gerchman, S. E., and Grollman, A. P. (2000) Substrate specificity and reaction mechanism of murine 8-oxoguanine-DNA glycosylase, J. Biol. Chem., 275, 28607–28617, doi: 10.1074/jbc. M002441200.

    Article  CAS  PubMed  Google Scholar 

  20. Bruner, S. D., Norman, D. P. G., and Verdine, G. L. (2000) Structural basis for recognition and repair of the endogenous mutagen 8-oxoguanine in DNA, Nature, 403, 859–866, doi: 10.1038/35002510.

    Article  CAS  PubMed  Google Scholar 

  21. Norman, D. P. G., Chung, S. J., and Verdine, G. L. (2003) Structural and biochemical exploration of a critical amino acid in human 8-oxoguanine glycosylase, Biochemistry, 42, 1564–1572, doi: 10.1021/bi026823d.

    Article  CAS  PubMed  Google Scholar 

  22. Norman, D. P. G., Bruner, S. D., and Verdine, G. L. (2001) Coupling of substrate recognition and catalysis by a human base-excision DNA repair protein, J. Am. Chem. Soc., 123, 359–360, doi: 10.1021/ja003144m.

    Article  CAS  PubMed  Google Scholar 

  23. Bjoras, M., Seeberg, E., Luna, L., Pearl, L. H., and Barrett, T. E. (2002) Reciprocal “flipping” underlies substrate recognition and catalytic activation by the human 8-oxoguanine DNA glycosylase, J. Mol. Biol., 317, 171–177, doi: 10.1006/jmbi.2002.5400.

    Article  CAS  PubMed  Google Scholar 

  24. Fromme, J. C., Bruner, S. D., Yang, W., Karplus, M., and Verdine, G. L. (2003) Product-assisted catalysis in base-excision DNA repair, Nat. Struct. Biol., 10, 204–211, doi: 10.1038/nsb902.

    Article  CAS  PubMed  Google Scholar 

  25. Chung, S. J., and Verdine, G. L. (2004) Structures of end products resulting from lesion processing by a DNA glyco-sylase/lyase, Chem. Biol., 11, 1643–1649, doi: 10.1016/j.chembiol.2004.09.014.

    Article  CAS  PubMed  Google Scholar 

  26. Banerjee, A., Yang, W., Karplus, M., and Verdine, G. L. (2005) Structure of a repair enzyme interrogating undamaged DNA elucidates recognition of damaged DNA, Nature, 434, 612–618, doi: 10.1038/nature03458.

    Article  CAS  PubMed  Google Scholar 

  27. Banerjee, A., and Verdine, G. L. (2006) A nucleobase lesion remodels the interaction of its normal neighbor in a DNA glycosylase complex, Proc. Natl. Acad. Sci. USA, 103, 15020–15025, doi: 10.1073/pnas.0603644103.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Radom, C. T., Banerjee, A., and Verdine, G. L. (2007) Structural characterization of human 8-oxoguanine DNA glycosylase variants bearing active site mutations, J. Biol. Chem., 282, 9182–9194, doi: 10.1074/jbc.M608989200.

    Article  CAS  PubMed  Google Scholar 

  29. Lee, S., Radom, C. T., and Verdine, G. L. (2008) Trapping and structural elucidation of a very advanced intermediate in the lesion-extrusion pathway of hOGG1, J. Am. Chem. Soc., 130, 7784–7785, doi: 10.1021/ja800821t.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Crenshaw, C. M., Nam, K., Oo, K., Kutchukian, P. S., Bowman, B. R., Karplus, M., and Verdine, G. L. (2012) Enforced presentation of an extrahelical guanine to the lesion recognition pocket of human 8-oxoguanine glycosy-lase, hOGG1, J. Biol. Chem., 287, 24916–24928, doi: 10.1074/jbc.M111.316497.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Li, H., Endutkin, A. V., Bergonzo, C., Fu, L., Grollman, A. P., Zharkov, D. O., and Simmerling, C. (2017) DNA deformation-coupled recognition of 8-oxoguanine: con-formational kinetic gating in human DNA glycosylase, J. Am. Chem. Soc., 139, 2682–2692, doi: 10.1021/jacs. 6b11433.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Lukina, M. V., Popov, A. V., Koval, V. V., Vorobjev, Y. N., Fedorova, O. S., and Zharkov, D. O. (2013) DNA damage processing by human 8-oxoguanine-DNA glycosylase mutants with the occluded active site, J. Biol. Chem., 288, 28936–28947, doi: 10.1074/jbc.M113.487322.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Kabsch, W. (1976) A solution for the best rotation to relate two sets of vectors, Acta Crystallogr. A, 32, 922–923, doi: 10.1107/S0567739476001873.

    Article  Google Scholar 

  34. Popov, A. V., and Vorobjev, Y. N. (2010) GUI-BioPASED program for molecular dynamics simulations of biopolymers with a graphical user interface, Mol. Biol., 44, 735–742, doi: 10.1134/S0026893310040217.

    CAS  Google Scholar 

  35. Case, D. A., Darden, T. A., Cheatham, T. E., III, Simmerling, C. L., Wang, J., et al. (2012) AMBER 12, University of California, San Francisco.

    Google Scholar 

  36. Perlow-Poehnelt, R. A., Zharkov, D. O., Grollman, A. P., and Broyde, S. (2004) Substrate discrimination by for-mamidopyrimidine-DNA glycosylase: distinguishing interactions within the active site, Biochemistry, 43, 16092–16105, doi: 10.1021/bi048747f.

    Article  CAS  PubMed  Google Scholar 

  37. Vorobjev, Y. N. (2011) Advances in implicit models of water solvent to compute conformational free energy and molecular dynamics of proteins at constant pH, Adv. Protein Chem. Struct. Biol., 85, 281–322, doi: 10.1016/B978-0-12-386485-7.00008-9.

    Article  CAS  PubMed  Google Scholar 

  38. Manning, G. S. (1978) The molecular theory of polyelec-trolyte solutions with applications to the electrostatic properties of polynucleotides, Q. Rev. Biophys., 11, 179–246, doi: 10.1017/s0033583500002031.

    Article  CAS  PubMed  Google Scholar 

  39. Ravishanker, G., Auffinger, P., Langley, D. R., Jayaram, B., Young, M. A., and Beveridge, D. L. (1997) Treatment of counterions in computer simulations of DNA, Rev. Comput. Chem., 11, 317–372, doi: 10.1002/9780470125885. ch6.

    CAS  Google Scholar 

  40. Popov, A. V., Vorobjev, Y. N., and Zharkov, D. O. (2013) MDTRA: a molecular dynamics trajectory analyzer with a graphical user interface, J. Comput. Chem., 34, 319–325, doi: 10.1002/jcc.23135.

    Article  CAS  PubMed  Google Scholar 

  41. Humphrey, W., Dalke, A., and Schulten, K. (1996) VMD: visual molecular dynamics, J. Mol. Graph., 14, 33–38, doi: 10.1016/0263-7855(96)00018-5.

    Article  CAS  PubMed  Google Scholar 

  42. Sayle, R. A., and Milner-White, E. J. (1995) RASMOL: biomolecular graphics for all, Trends Biochem. Sci., 20, 374–376, doi: 10.1016/S0968-0004(00)89080-5.

    Article  CAS  PubMed  Google Scholar 

  43. Yang, W., Bitetti-Putzer, R., and Karplus, M. (2004) Free energy simulations: use of reverse cumulative averaging to determine the equilibrated region and the time required for convergence, J. Chem. Phys., 120, 2618–2628, doi: 10.1063/1.1638996.

    Article  CAS  PubMed  Google Scholar 

  44. Letunic, I., and Bork, P. (2016) Interactive tree of life (iTOL) v3: an online tool for the display and annotation of phylogenetic and other trees, Nucleic Acids Res., 44, W242–W245, doi: 10.1093/nar/gkw290.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Sambrook, J., and Russell, D. W. (2001) Molecular Cloning: a Laboratory Manual, 3rd Edn., Cold Spring Harbor Laboratory Press, Cold Spring Harbor.

    Google Scholar 

  46. Sidorenko, V. S., Nevinsky, G. A., and Zharkov, D. O. (2007) Mechanism of interaction between human 8-oxoguanine-DNA glycosylase and AP endonuclease, DNA Repair, 6, 317–328, doi: 10.1016/j.dnarep.2006.10.022.

    Article  CAS  PubMed  Google Scholar 

  47. Sidorenko, V. S., Mechetin, G. V., Nevinsky, G. A., and Zharkov, D. O. (2008) Ionic strength and magnesium affect the specificity of Escherichia coli and human 8-oxogua-nine-DNA glycosylases, FEBS J., 275, 3747–3760, doi: 10.1111/j.1742-4658.2008.06521.x.

    Article  CAS  PubMed  Google Scholar 

  48. Anderson, P. C., and Daggett, V. (2009) The R46Q, R131Q and R154H polymorphs of human DNA glycosylase/ß-lyase hOgg1 severely distort the active site and DNA recognition site but do not cause unfolding, J. Am. Chem. Soc., 131, 9506–9515, doi: 10.1021/ja809726e.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Sowlati-Hashjin, S., and Wetmore, S. D. (2018) Structural insight into the discrimination between 8-oxoguanine gly-cosidic conformers by DNA repair enzymes: a molecular dynamics study of human oxoguanine glycosylase 1 and formamidopyrimidine-DNA glycosylase, Biochemistry, 57, 1144–1154, doi: 10.1021/acs.biochem.7b01292.

    Article  CAS  PubMed  Google Scholar 

  50. Bjoras, M., Luna, L., Johnsen, B., Hoff, E., Haug, T., Rognes, T., and Seeberg, E. (1997) Opposite base-dependent reactions of a human base excision repair enzyme on DNA containing 7,8-dihydro-8-oxoguanine and abasic sites, EMBO J., 16, 6314–6322, doi: 10.1093/emboj/16.20.6314.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Kuznetsov, N. A., Koval, V. V., Zharkov, D. O., Nevinsky, G. A., Douglas, K. T., and Fedorova, O. S. (2005) Kinetics of substrate recognition and cleavage by human 8-oxogua-nine-DNA glycosylase, Nucleic Acids Res., 33, 3919–3931, doi: 10.1093/nar/gki694.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Zharkov, D. O., Golan, G., Gilboa, R., Fernandes, A. S., Gerchman, S. E., Kycia, J. H., Rieger, R. A., Grollman, A. P., and Shoham, G. (2002) Structural analysis of an Escherichia coli endonuclease VIII covalent reaction intermediate, EMBO J., 21, 789–800, doi: 10.1093/emboj/21.4.789.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Li, H., Endutkin, A. V., Bergonzo, C., Campbell, A. J., de los Santos, C., Grollman, A., Zharkov, D. O., and Simmerling, C. (2016) A dynamic checkpoint in oxidative lesion discrimination by formamidopyrimidine-DNA gly-cosylase, Nucleic Acids Res., 44, 683–694, doi: 10.1093/nar/gkv1092.

    Article  PubMed  CAS  Google Scholar 

  54. Sowlati-Hashjin, S., and Wetmore, S. D. (2014) Computational investigation of glycosylase and ß-lyase activity facilitated by proline: applications to FPG and comparisons to hOgg1, J. Phys. Chem. B, 118, 14566–14577, doi: 10.1021/jp507783d.

    Article  CAS  PubMed  Google Scholar 

  55. Popov, A. V., Endutkin, A. V., Vorobjev, Y. N., and Zharkov, D. O. (2017) Molecular dynamics simulation of the opposite-base preference and interactions in the active site of formamidopyrimidine-DNA glycosylase, BMC Struct. Biol., 17, 5, doi: 10.1186/s12900-017-0075-y.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

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Acknowledgements

The calculations were performed at the supercomputer NKS-30T cluster of the Siberian Supercomputer Center, Siberian Branch of the Russian Academy of Sciences.

Funding

The study was supported by the Russian Science Foundation (project 18-74-00052; simulation, enzyme kinetics), Program for Fundamental Scientific Research of the State Academies of Sciences for 2013-2020 (project AAAA-A17-117020210023-1; oligonu-cleotide synthesis, protein purification), and Ministry of Education and Science of the Russian Federation (project 6.5773.2017/VU, data analysis).

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Authors and Affiliations

  1. Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, 630090, Novosibirsk, Russia

    A. V. Popov, A. V. Yudkina, Yu. N. Vorobjev & D. O. Zharkov

  2. Novosibirsk State University, 630090, Novosibirsk, Russia

    A. V. Yudkina & D. O. Zharkov

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  1. A. V. Popov
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  2. A. V. Yudkina
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  3. Yu. N. Vorobjev
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  4. D. O. Zharkov
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Correspondence to A. V. Popov or D. O. Zharkov.

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Compliance with ethical standard. This article does not contain any studies involving animals or human participants performed by any of the authors.

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Conflict of interest. The authors declare no conflict of interest.

Russian Text © The Author(s), 2020, published in Biokhimiya, 2020, Vol. 85, No. 2, pp. 225-238.

Originally published in Biochemistry (Moscow) On-Line Papers in Press, as Manuscript BM19-116, January 6, 2020.

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Popov, A.V., Yudkina, A.V., Vorobjev, Y.N. et al. Catalytically Competent Conformation of the Active Site of Human 8-Oxoguanine-DNA Glycosylase. Biochemistry Moscow 85, 192–204 (2020). https://doi.org/10.1134/S0006297920020066

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