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. 1974 Jan;71(1):205–208. doi: 10.1073/pnas.71.1.205

Metabolic Regulation of Cytoplasmic DNA Synthesis

John J Byrnes 1,2, Kathleen M Downey 1,2, Antero G So 1,2
PMCID: PMC387966  PMID: 4521051

Abstract

The regulation of cytoplasmic DNA synthesis by the metabolites ATP and citrate has been demonstrated. Other ribonucleoside and deoxyribonucleoside triphosphates as well as α,β-methylene- and β,γ-methylene-ATP and α,β-methylene-ADP are able to partially substitute for ATP in stimulating the rate of DNA synthesis with the cytoplasmic DNA polymerase (DNA nucleotidyltransferase, EC 2.7.7.7) from bone marrow. The fact that the methylene analogs of ATP and ADP are effective in stimulating DNA synthesis indicates that the mechanism of stimulation does not involve ATP hydrolysis.

The nucleotide activators have been shown by kinetic analysis to affect the Vmax of the enzyme and not the apparent Kms for the substrates. The curve that results when the rate of DNA synthesis is plotted as a function of ATP concentration is sigmoidal, suggesting that more than one site on the enzyme interacts with ATP and that these sites are acting cooperatively. The concentration of ATP required for maximal velocity is dependent on the Mn++ concentration. At pH 7.0 maximal activity is obtained when the molar ratio of ATP to Mn++ is 1.6:1. When either ATP or Mn++ is present in relative excess, DNA synthesis is inhibited.

The mechanism of ATP activation has been shown to be associated with an alteration in the sedimentation behavior of the DNA polymerase. In the presence of ATP, there is an increase in the fraction of the enzyme that sediments at 8 S with a corresponding decrease in the 11.6S enzyme fraction. Thus, ATP activation corresponds to the dissociation of an 11.6S dimer into 8S monomers.

In addition to ATP and other nucleotides, citrate also stimulates DNA synthesis. At present it is not clear whether the stimulatory effects of ATP and citrate are due to their ability to chelate Mn++, which is inhibitory at high concentrations, or whether an ATP-Mn++ or citrate-Mn++ complex is the activator.

Keywords: cytoplasmic DNA polymerase, enzyme activation by ATP, dimer dissociation

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Selected References

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

  1. APOSHIAN H. V., KORNBERG A. Enzymatic synthesis of deoxyribonucleic acid. IX. The polymerase formed after T2 bacteriophage infection of Escherichia coli: a new enzyme. J Biol Chem. 1962 Feb;237:519–525. [PubMed] [Google Scholar]
  2. Blakley R. L., Vitols E. The control of nucleotide biosynthesis. Annu Rev Biochem. 1968;37:201–224. doi: 10.1146/annurev.bi.37.070168.001221. [DOI] [PubMed] [Google Scholar]
  3. Byrnes J. J., Downey K. M., So A. G. Bone marrow cytoplasmic deoxyribonucleic acid polymerase. Variation of pH and ionic environment as a possible control mechanism. Biochemistry. 1973 Oct 23;12(22):4378–4384. doi: 10.1021/bi00746a013. [DOI] [PubMed] [Google Scholar]
  4. Elford H. L. Functional regulation of mammalian ribonucleotide reductase. Adv Enzyme Regul. 1972;10:19–38. doi: 10.1016/0065-2571(72)90004-0. [DOI] [PubMed] [Google Scholar]
  5. Mordoh J., Hirota Y., Jacob F. On the process of cellular division in Escherichia coli. V. Incorporation of deoxynucleoside triphosphates by DNA thermosensitive mutants of Escherichia coli also lacking DNA polymerase activity. Proc Natl Acad Sci U S A. 1970 Oct;67(2):773–778. doi: 10.1073/pnas.67.2.773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Moses R. E., Richardson C. C. Replication and repair of DNA in cells of Escherichia coli treated with toluene. Proc Natl Acad Sci U S A. 1970 Oct;67(2):674–681. doi: 10.1073/pnas.67.2.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Randle P. J., Denton R. M., England P. J. Citrate as a metabolic regulator in muscle and adipose tissue. Biochem Soc Symp. 1968;27:87–103. [PubMed] [Google Scholar]
  8. Schekman R., Wickner W., Westergaard O., Brutlag D., Geider K., Bertsch L. L., Kornberg A. Initiation of DNA synthesis: synthesis of phiX174 replicative form requires RNA synthesis resistant to rifampicin. Proc Natl Acad Sci U S A. 1972 Sep;69(9):2691–2695. doi: 10.1073/pnas.69.9.2691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Stadtman E. R. Allosteric regulation of enzyme activity. Adv Enzymol Relat Areas Mol Biol. 1966;28:41–154. doi: 10.1002/9780470122730.ch2. [DOI] [PubMed] [Google Scholar]
  10. Söderhäll S. S., Larsson A., Skoog K. L. Deoxyribonucleotide pools during liver regeneration. Eur J Biochem. 1973 Feb 15;33(1):36–39. doi: 10.1111/j.1432-1033.1973.tb02651.x. [DOI] [PubMed] [Google Scholar]
  11. Vosberg H. P., Hoffmann-Berling H. DNA synthesis in nucleotide-permeable Escherichia coli cells. I. Preparation and properties of ether-treated cells. J Mol Biol. 1971 Jun 28;58(3):739–753. doi: 10.1016/0022-2836(71)90037-4. [DOI] [PubMed] [Google Scholar]

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