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. 1977 Oct;4(10):3589–3597. doi: 10.1093/nar/4.10.3589

The effect of magnesium and manganese ions on the structure and template activity for reverse transcriptase of polyribocytidylate and its 2'-0-methyl derivative.

N C Vamvakopoulos, J N Vournakis, S L Marcus
PMCID: PMC342675  PMID: 73165

Abstract

The secondary structure of the hydrogen bonded hybrids polycytidylate-oligodeoxguanylate (poly(rC)-(dG)12-18 and poly (2'-oMe) cytidylate-oligodeoxyguanylate (poly (rCm)-(dG)12-18 was studied at several magnesium and manganese ion concentrations. These hybrids are effective template-primer complexes for the synthesis of poly(dG) by avian myeloblastosis virus (AMV) DNA polymerase under disparate ionic conditions. Circular dichroism spectra and thermal melting data were obtained as a function of ion concentration, including conditions that allow optimum rates of poly (dG) synthesis by each complex. These studies demonstrate that both hybrids can change conformation and stability depending on their ionic environment. Comparison of enzyme activity and physical data suggest that the polymerase recognizes particular secondary structure features. Changes in the activity of the AMV polymerase can be induced by varying the Mg++ and Mn++ concentrations alone and in combination. These variations in enzyme activity are correlated with observed changes in the base-stacking alignment of the synthetic template primers. The ions, therefore, seem to affect enzyme activity by altering the conformation of the polnucleotide complexes.

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Selected References

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

  1. Auld D. S., Kawaguchi H., Livingston D. M., Vallee B. L. RNA-dependent DNA polymerase (reverse transcriptase) from avian myeloblastosis virus: a zinc metalloenzyme. Proc Natl Acad Sci U S A. 1974 May;71(5):2091–2095. doi: 10.1073/pnas.71.5.2091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baltimore D., Smoler D. Primer requirement and template specificity of the DNA polymerase of RNA tumor viruses. Proc Natl Acad Sci U S A. 1971 Jul;68(7):1507–1511. doi: 10.1073/pnas.68.7.1507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gerard G. F., Loewenstein P. M., Green M., Rottman F. Detection of reverse transcriptase in human breast tumours with poly(Cm)-oligo(dG). Nature. 1975 Jul 10;256(5513):140–143. doi: 10.1038/256140a0. [DOI] [PubMed] [Google Scholar]
  4. Gerard G. F., Rottman F., Green M. Poly(2'-O-methylcytidylate)-oligodeoxyguanylate as a template for the ribonucleic acid directed deoxyribonucleic acid polymerase in ribonucleic acid tumor virus particles and a specific probe for the ribonucleic acid directed enzyme in transformed murine cells. Biochemistry. 1974 Apr 9;13(8):1632–1641. doi: 10.1021/bi00705a012. [DOI] [PubMed] [Google Scholar]
  5. Gerard G. F., Rottman F., Green M. Poly(2'-O-methylcytidylate)-oligodeoxyguanylate as a template for the ribonucleic acid directed deoxyribonucleic acid polymerase in ribonucleic acid tumor virus particles and a specific probe for the ribonucleic acid directed enzyme in transformed murine cells. Biochemistry. 1974 Apr 9;13(8):1632–1641. doi: 10.1021/bi00705a012. [DOI] [PubMed] [Google Scholar]
  6. Goodman N. C., Spiegelman S. Distinguishing reverse transcriptase of an RNA tumor virus from other known DNA polymerases. Proc Natl Acad Sci U S A. 1971 Sep;68(9):2203–2206. doi: 10.1073/pnas.68.9.2203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Green M., Gerard G. F. RNA-directed DNA polymerase--properties and functions in oncogenic RNA viruses and cells. Prog Nucleic Acid Res Mol Biol. 1974;14(0):187–334. [PubMed] [Google Scholar]
  8. Marcus S. L., Modak M. J. Observations on template-specific conditions for DNA synthesis by avian myeloblastosis virus DNA polymerase. Nucleic Acids Res. 1976 Jun;3(6):1473–1486. doi: 10.1093/nar/3.6.1473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Marcus S. L., Sarkar N. H., Modak M. J. Purification and properties of murine mammary tumor virus DNA polymerase. Virology. 1976 May;71(1):242–254. doi: 10.1016/0042-6822(76)90109-4. [DOI] [PubMed] [Google Scholar]
  10. Mikke R., Zmudzka B. Oligo(dG)12-18 aggregates result in non-homogeneity of oligo(dG)12-18.poly(C) type primer-template. Nucleic Acids Res. 1977 Apr;4(4):1111–1122. doi: 10.1093/nar/4.4.1111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Modak M. J., Marcus S. L. Purification and properties of Rauscher leukemia virus DNA polymerase and selective inhibition of mammalian viral reverse transcriptase by inorganic phosphate. J Biol Chem. 1977 Jan 10;252(1):11–19. [PubMed] [Google Scholar]
  12. Tamblyn T. M., Wells R. D. Comparative ability of RNA and DNA to prime DNA synthesis in vitro: role of sequence, sugar, and structure of template-primer. Biochemistry. 1975 Apr 8;14(7):1412–1425. doi: 10.1021/bi00678a011. [DOI] [PubMed] [Google Scholar]
  13. Walter A., Luck G. Interactions of Hg(II) ions with DNA as revealed by CD measurements. Nucleic Acids Res. 1977 Mar;4(3):539–550. doi: 10.1093/nar/4.3.539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Zimmer C., Luck G., Holy A. CD studies on the conformation of oligonucleotides complexed with divalent metal ions: interaction of Zn2+ with guanine favours syn conformation. Nucleic Acids Res. 1976 Oct;3(10):2757–2770. doi: 10.1093/nar/3.10.2757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Zmudzka B., Janion C., Shugar D. Poly 2'-O-methylcytidylic acid and the role of the 2'-hydroxyl in polynucleotide structure. Biochem Biophys Res Commun. 1969 Dec 4;37(6):895–901. doi: 10.1016/0006-291x(69)90215-0. [DOI] [PubMed] [Google Scholar]

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