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. 1989 Apr;86(7):2147–2151. doi: 10.1073/pnas.86.7.2147

Oxygen-sensitive ribonucleoside triphosphate reductase is present in anaerobic Escherichia coli.

M Fontecave 1, R Eliasson 1, P Reichard 1
PMCID: PMC286868  PMID: 2648390

Abstract

Ribonucleoside diphosphate reductase from Escherichia coli and mammalian cells provides the deoxyribonucleoside triphosphates for DNA synthesis. The active enzyme contains a tyrosyl free radical whose formation requires oxygen. Earlier genetic evidence suggested that the enzyme is not required for anaerobic growth of E. coli, implicating the activity of a different enzyme or enzyme system for deoxyribonucleotide synthesis in the absence of oxygen. We now conclude from isotope incorporation experiments that E. coli during anaerobiosis obtains its deoxyribonucleotides by reduction of ribonucleotides. Extracts from anaerobically grown bacteria contain a different enzyme activity capable of reducing CTP to dCTP. To obtain an active enzyme, strict anaerobiosis must be maintained during extract preparation and during assay of the enzyme. The reaction is stimulated by NADPH, Mg2+, and ATP. Inhibition by deoxyribonucleoside triphosphates suggests that the anaerobic enzyme has allosteric properties. Antibodies raised against the aerobic enzyme do not inhibit the new activity, and hydroxyurea, a potent scavenger of the tyrosyl radical of the aerobic enzyme, only weakly inhibits the anaerobic enzyme. The anaerobic enzyme has interesting evolutionary aspects since it might reflect on an activity that in the absence of oxygen made possible the transition from an "RNA world" into a "DNA world."

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Selected References

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

  1. Barlow T., Eliasson R., Platz A., Reichard P., Sjöberg B. M. Enzymic modification of a tyrosine residue to a stable free radical in ribonucleotide reductase. Proc Natl Acad Sci U S A. 1983 Mar;80(6):1492–1495. doi: 10.1073/pnas.80.6.1492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barlow T. Evidence for a new ribonucleotide reductase in anaerobic E. coli. Biochem Biophys Res Commun. 1988 Sep 15;155(2):747–753. doi: 10.1016/s0006-291x(88)80558-8. [DOI] [PubMed] [Google Scholar]
  3. Bianchi V., Pontis E., Reichard P. Regulation of pyrimidine deoxyribonucleotide metabolism by substrate cycles in dCMP deaminase-deficient V79 hamster cells. Mol Cell Biol. 1987 Dec;7(12):4218–4224. doi: 10.1128/mcb.7.12.4218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Ehrenberg A., Reichard P. Electron spin resonance of the iron-containing protein B2 from ribonucleotide reductase. J Biol Chem. 1972 Jun 10;247(11):3485–3488. [PubMed] [Google Scholar]
  6. Fontecave M., Eliasson R., Reichard P. NAD(P)H:flavin oxidoreductase of Escherichia coli. A ferric iron reductase participating in the generation of the free radical of ribonucleotide reductase. J Biol Chem. 1987 Sep 5;262(25):12325–12331. [PubMed] [Google Scholar]
  7. Gräslund A., Ehrenberg A., Thelander L. Characterization of the free radical of mammalian ribonucleotide reductase. J Biol Chem. 1982 May 25;257(10):5711–5715. [PubMed] [Google Scholar]
  8. Hantke K. Characterization of an iron sensitive Mud1 mutant in E. coli lacking the ribonucleotide reductase subunit B2. Arch Microbiol. 1988;149(4):344–349. doi: 10.1007/BF00411654. [DOI] [PubMed] [Google Scholar]
  9. Jamison C. S., Adler H. I. Mutations in Escherichia coli that effect sensitivity to oxygen. J Bacteriol. 1987 Nov;169(11):5087–5094. doi: 10.1128/jb.169.11.5087-5094.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jeter R. M., Olivera B. M., Roth J. R. Salmonella typhimurium synthesizes cobalamin (vitamin B12) de novo under anaerobic growth conditions. J Bacteriol. 1984 Jul;159(1):206–213. doi: 10.1128/jb.159.1.206-213.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Karlström O., Larsson A. Significance of ribonucleotide reduction in the biosynthesis of deoxyribonucleotides in Escherichia coli. Eur J Biochem. 1967 Dec;3(2):164–170. doi: 10.1111/j.1432-1033.1967.tb19512.x. [DOI] [PubMed] [Google Scholar]
  12. Khym J. X. An analytical system for rapid separation of tissue nucleotides at low pressures on conventional anion exchangers. Clin Chem. 1975 Aug;21(9):1245–1252. [PubMed] [Google Scholar]
  13. RACKER E. Enzymatic synthesis and breakdown of desoxyribose phosphate. J Biol Chem. 1952 May;196(1):347–365. [PubMed] [Google Scholar]
  14. REICHARD P. Enzymatic synthesis of deoxyribonucleotides. I. Formation of deoxycytidine diphosphate from cytidine diphosphate with enzymes from Escherichia coli. J Biol Chem. 1962 Nov;237:3513–3519. [PubMed] [Google Scholar]
  15. Reichard P. Interactions between deoxyribonucleotide and DNA synthesis. Annu Rev Biochem. 1988;57:349–374. doi: 10.1146/annurev.bi.57.070188.002025. [DOI] [PubMed] [Google Scholar]
  16. Saeki T., Hori M., Umezawa H. Pyruvate kinase of Escherichia coli. Its role in supplying nucleoside triphosphates in cells under anaerobic conditions. J Biochem. 1974 Sep;76(3):631–637. doi: 10.1093/oxfordjournals.jbchem.a130607. [DOI] [PubMed] [Google Scholar]
  17. Sjöberg B. M., Hahne S., Mathews C. Z., Mathews C. K., Rand K. N., Gait M. J. The bacteriophage T4 gene for the small subunit of ribonucleotide reductase contains an intron. EMBO J. 1986 Aug;5(8):2031–2036. doi: 10.1002/j.1460-2075.1986.tb04460.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Sjöberg B. M., Reichard P., Gräslund A., Ehrenberg A. The tyrosine free radical in ribonucleotide reductase from Escherichia coli. J Biol Chem. 1978 Oct 10;253(19):6863–6865. [PubMed] [Google Scholar]
  19. Thelander L., Eriksson S., Akerman M. Ribonucleotide reductase from calf thymus. Separation of the enzyme into two nonidentical subunits, proteins M1 and M2. J Biol Chem. 1980 Aug 10;255(15):7426–7432. [PubMed] [Google Scholar]
  20. Thelander L., Reichard P. Reduction of ribonucleotides. Annu Rev Biochem. 1979;48:133–158. doi: 10.1146/annurev.bi.48.070179.001025. [DOI] [PubMed] [Google Scholar]
  21. WEHRLI W. E., VERHEYDEN D. L., MOFFATT J. G. DISMUTATION REACTIONS OF NUCLEOSIDE POLYPHOSPHATES. II. SPECIFIC CHEMICAL SYNTHESES OF ALPHA-, BETA-, AND GAMMA-P32-NUCLEOSIDE 5'-TRIPHOSPHATES. J Am Chem Soc. 1965 May 20;87:2265–2277. doi: 10.1021/ja01088a028. [DOI] [PubMed] [Google Scholar]

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