AIMS Biophysics, 2018, 5(2): 125-143. doi: 10.3934/biophy.2018.2.125.

Research article

Export file:

Format

  • RIS(for EndNote,Reference Manager,ProCite)
  • BibTex
  • Text

Content

  • Citation Only
  • Citation and Abstract

The effect of different divalent cations on the kinetics and fidelity of Bacillus stearothermophilus DNA polymerase

1 Centre for Structural and Functional Genomics, Concordia University, Montreal, Canada
2 Yale University, 333 Cedar St., SHM C-E14, New Haven, CT, 06520, USA

Although Mg2+ is the metal ion that functions as the cofactor for DNA polymerases (DNA pols) in vivo, Mn2+ can also serve in this capacity but it reduces base discrimination. Metal ions aside from Mg2+ or Mn2+ can act as cofactors for some DNA pols but not for others. Here we report on the ability of several divalent metal ions to substitute for Mg2+ or Mn2+ with BST DNA polymerase (BST pol), an A family DNA pol. We selected the metal ions based on whether they had previously been shown to be effective with other DNA pols. We found that Co2+ and Cd2+ were the only cations tested that could replace Mg2+ or Mn2+. When Co2+ was substituted for Mg2+, the incorporation efficiency for correct dNTPs increased 6-fold but for incorrect dNTPs there was a decrease which depended on the incoming dNTP. With Mn2+, base selectivity was impaired compared to Co2+ and Cd2+. In addition, Co2+ and Mn2+ helped BST pol to catalyze primer-extension past a mismatch. Finally both Co2+ and Mn2+ enhanced ground-state binding of both correct and incorrect dNTPs to BST pol: Dideoxy terminated primer-template complexes.
  Figure/Table
  Supplementary
  Article Metrics

Keywords divalent cations; base selectivity; BST pol; kinetics

Citation: Ashwani Kumar Vashishtha, William H. Konigsberg. The effect of different divalent cations on the kinetics and fidelity of Bacillus stearothermophilus DNA polymerase. AIMS Biophysics, 2018, 5(2): 125-143. doi: 10.3934/biophy.2018.2.125

References

  • 1. Drake JW (1969) Comparative rates of spontaneous mutation. Nature 221: 1132.    
  • 2. Steitz TA, Steitz JA (1993) A general two-metal-ion mechanism for catalytic RNA. P Natl Acad Sci USA 90: 6498–6502.    
  • 3. Vaisman A, Ling H, Woodgate R, et al. (2005) Fidelity of Dpo4: Effect of metal ions, nucleotide selection and pyrophosphorolysis. EMBO J 24: 2957–2967.    
  • 4. Pelletier H, Sawaya MR, Wolfle W, et al. (1996) A structural basis for metal ion mutagenicity and nucleotide selectivity in human DNA polymerase beta. Biochemistry 35: 12762–12777.    
  • 5. Irimia A, Loukachevitch LV, Eoff ARL, et al. (2010) Metal-ion dependence of the active-site conformation of the translesion DNA polymerase Dpo4 from Sulfolobus solfataricus. Acta Crystallogr 66: 1013–1018.
  • 6. Irimia A, Zang H, Loukachevitch LV, et al. (2006) Calcium is a cofactor of polymerization but inhibits pyrophosphorolysis by the Sulfolobus solfataricus DNA polymerase Dpo4. Biochemistry 45: 5949–5956.
  • 7. Xia S, Wang M, Blaha G, et al. (2011) Structural insights into complete metal ion coordination from ternary complexes of B family RB69 DNA polymerase. Biochemistry 50: 9114–9124.    
  • 8. Nakamura T, Zhao Y, Yamagata Y, et al. (2012) Watching DNA polymerase eta make a phosphodiester bond. Nature 487: 196–201.    
  • 9. Sirover MA, Dube DK, Loeb LA (1979) On the fidelity of DNA replication. Metal activation of Escherichia coli DNA polymerase I. J Biol Chem 254: 107–111.
  • 10. Sirover MA, Loeb LA (1977) On the fidelity of DNA replication. Effect of metal activators during synthesis with avian myeloblastosis virus DNA polymerase. J Biol Chem 252: 3605–3610.
  • 11. Miyaki M, Murata I, Osabe M, et al. (1977) Effect of metal cations on misincorporation by E. coli DNA polymerases. Biochem Bioph Res Co 77: 854–860.    
  • 12. Goodman MF, Keener S, Guidotti S, et al. (1983) On the enzymatic basis for mutagenesis by manganese. J Biol Chem 258: 3469–3475.
  • 13. Snow ET, Xu LS, Kinney PL (1993) Effects of nickel ions on polymerase activity and fidelity during DNA replication in vitro. Chem-Biol Interact 88: 155–173.
  • 14. Vashishtha AK, Wang J, Konigsberg WH (2016) Different Divalent Cations Alter the Kinetics and Fidelity of DNA Polymerases. J Biol Chem 291: 20869–20875.    
  • 15. Sirover MA, Loeb LA (1976) Infidelity of DNA synthesis in vitro: Screening for potential metal mutagens or carcinogens. Science 194: 1434–1436.    
  • 16. Hartwig A, Asmuss M, Ehleben I, et al. (2002) Interference by toxic metal ions with DNA repair processes and cell cycle control: Molecular mechanisms. Environ Health Persp 110: 797–799.
  • 17. Vashishtha AK, Konigsberg WH (2016) Effect of Different Divalent Cations on the Kinetics and Fidelity of RB69 DNA Polymerase. Biochemistry 55: 2661–2670.    
  • 18. Johnson SJ, Taylor JS, Beese LS (2003) Processive DNA synthesis observed in a polymerase crystal suggests a mechanism for the prevention of frameshift mutations. P Natl Acad Sci USA 100: 3895–3900.    
  • 19. Johnson SJ, Beese LS (2004) Structures of mismatch replication errors observed in a DNA polymerase. Cell 116: 803–816.    
  • 20. Kuchta RD, Mizrahi V, Benkovic PA, et al. (1987) Kinetic mechanism of DNA polymerase I (Klenow). Biochemistry 26: 8410–8417.
  • 21. Kiefer JR, Mao C, Hansen CJ, et al. (1997) Crystal structure of a thermostable Bacillus DNA polymerase I large fragment at 2.1 A resolution. Structure 5: 95–108.
  • 22. Hays H, Berdis AJ (2002) Manganese substantially alters the dynamics of translesion DNA synthesis. Biochemistry 41: 4771–4778.    
  • 23. Wang TS, Sedwick WD, Korn D (1974) Nuclear deoxyribonucleic acid polymerase. Purification and properties of the homogeneous enzyme from human KB cells. J Biol Chem 249: 841–850.
  • 24. Seal G, Shearman CW, Loeb LA (1979) On the fidelity of DNA replication. Studies with human placenta DNA polymerases. J Biol Chem 254: 5229–5237.
  • 25. Villani G, Tanguy LGN, Wasungu L, et al. (2002) Effect of manganese on in vitro replication of damaged DNA catalyzed by the herpes simplex virus type-1 DNA polymerase. Nucleic Acids Res 30: 3323–3332.    
  • 26. Wang W, Hellinga HW, Beese LS (2011) Structural evidence for the rare tautomer hypothesis of spontaneous mutagenesis. P Natl Acad Sci USA 108: 17644–17648.    
  • 27. Yu SL, Lee SK, Johnson RE, et al. (2003) The stalling of transcription at abasic sites is highly mutagenic, Mol Cell Biol 23: 382–388.
  • 28. Cowna JA (2002) Structural and catalytic chemistry of magnesium-dependent enzymes. Biometals 15: 225–235.    
  • 29. Bock CW, Katz AK, Markham GD, et al. (1999) Manganese as a Replacement for Magnesium and Zinc:  Functional Comparison of the Divalent Ions. J Am Chem Soc 121: 7360–7372.    
  • 30. Chin YE, Snow ET, Cohen MD, et al. (1994) The effect of divalent nickel (Ni2+) on in vitro DNA replication by DNA polymerase alpha. Cancer Res 54: 2337–2341.
  • 31. Xia S, Vashishtha A, Bulkley D, et al. (2012) Contribution of partial charge interactions and base stacking to the efficiency of primer extension at and beyond abasic sites in DNA. Biochemistry 51: 4922−4931.
  • 32. Vashishtha AK, Kuchta RD (2015) Polymerase and exonuclease activities in herpes simplex virus type 1 DNA polymerase are not highly coordinated. Biochemistry 54: 240−249.
  • 33. Vashishtha AK, Kuchta RD (2016) Effects of Acyclovir, Foscarnet and Ribonucleotides on Herpes Simplex Virus-1 DNA Polymerase: Mechanistic Insights and a Novel Mechanism for preventing Stable Incorporation of Ribonucleotides into DNA. Biochemistry 55: 1168−1177.

 

This article has been cited by

  • 1. Ashwani Kumar Vashishtha, William H. Konigsberg, Effect of Different Divalent Cations on the Kinetics and Fidelity of DNA Polymerases, AIMS Biophysics, 2018, 5, 4, 272, 10.3934/biophy.2018.4.272
  • 2. Max E. Gottesman, Maxim Chudaev, Arkady Mustaev, Key features of magnesium that underpin its role as the major ion for electrophilic biocatalysis, The FEBS Journal, 2020, 10.1111/febs.15318

Reader Comments

your name: *   your email: *  

© 2018 the Author(s), licensee AIMS Press. This is an open access article distributed under the terms of the Creative Commons Attribution Licese (http://creativecommons.org/licenses/by/4.0)

Download full text in PDF

Export Citation

Copyright © AIMS Press All Rights Reserved