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. 1970 Sep;103(3):622–633. doi: 10.1128/jb.103.3.622-633.1970

Studies on the Pathway of Incorporation of 2-Aminopurine into the Deoxyribonucleic Acid of Escherichia coli1

Eleanor G Rogan a,2, Maurice J Bessman a
PMCID: PMC248136  PMID: 4919986

Abstract

A pathway for the incorporation of 2-aminopurine into deoxyribonucleic acid (DNA) was studied in cell-free extracts of Escherichia coli. It was demonstrated that the free base can be converted to the deoxynucleoside, and that the deoxynucleotide can be phosphorylated to the di- and triphosphates and then incorporated into the DNA. From a consideration of the individual reactions in crude extracts, it is likely that the rate-limiting step in this pathway is the formation of the deoxynucleotide. Of especial interest is the observation that 2-aminopurine may be viewed as an analogue of either guanine or adenine, depending on which enzymatic step is being considered. On the one hand, it resembles guanine in that it is specifically converted from the mono- to the diphosphate by guanylate kinase and not by adenylate kinase. On the other hand, it replaces adenine rather than guanine in the DNA synthesized with purified DNA polymerases. E. coli DNA polymerase utilizes aminopurine deoxynucleoside triphosphate as a substrate for DNA synthesis much better than does purified phage T5-induced DNA polymerase and is also much less inhibited by this analogue than the T5 enzyme. These experiments in vitro correlate with known differential effects of 2-aminopurine on E. coli and phage in vivo.

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Selected References

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

  1. AMES B. N., DUBIN D. T. The role of polyamines in the neutralization of bacteriophage deoxyribonucleic acid. J Biol Chem. 1960 Mar;235:769–775. [PubMed] [Google Scholar]
  2. Abrams R., Edmonds M., Libenson L. Deoxyribosyl exchange activity associated with nucleoside phosphorylase. Biochem Biophys Res Commun. 1965 Jul 26;20(3):310–314. doi: 10.1016/0006-291x(65)90365-7. [DOI] [PubMed] [Google Scholar]
  3. BELLO L. J., BESSMAN M. J. THE ENZYMOLOGY OF VIRUS-INFECTED BACTERIA. V. PHOSPHORYLATION OF HYDROXYMETHYLDEOXYCYTIDINE DIPHOSPHATE AND DEOXYTHYMIDINE DIPHOSPHATE IN NORMAL AND BACTERIOPHAGE-INFECTED ESCHERICHIA COLI. Biochim Biophys Acta. 1963 Aug 20;72:647–650. doi: 10.1016/0006-3002(63)90293-2. [DOI] [PubMed] [Google Scholar]
  4. BELLO L. J., BESSMAN M. J. The enzymology of virus-infected bacteria. IV. Purification and properties of the deoxynucleotide kinase induced by bacteriophage T2. J Biol Chem. 1963 May;238:1777–1787. [PubMed] [Google Scholar]
  5. BESSMAN M. J., HERRIOTT S. T., ORR M. J. THE ENZYMOLOGY OF VIRUS-INFECTED BACTERIA. VI. PURIFICATION AND PROPERTIES OF THE DEOXYNUCLEOTIDE KINASE INDUCED BY BACTERIOPHAGE T5. J Biol Chem. 1965 Jan;240:439–445. [PubMed] [Google Scholar]
  6. BJORK W. Purification of phosphodiesterase from Bothrops atrox venom, with special consideration of the elimination of monophosphatases. J Biol Chem. 1963 Jul;238:2487–2490. [PubMed] [Google Scholar]
  7. BURTON K. A study of the conditions and mechanism of the diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem J. 1956 Feb;62(2):315–323. doi: 10.1042/bj0620315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bessman M. J., Lehman I. R., Adler J., Zimmerman S. B., Simms E. S., Kornberg A. ENZYMATIC SYNTHESIS OF DEOXYRIBONUCLEIC ACID. III. THE INCORPORATION OF PYRIMIDINE AND PURINE ANALOGUES INTO DEOXYRIBONUCLEIC ACID. Proc Natl Acad Sci U S A. 1958 Jul 15;44(7):633–640. doi: 10.1073/pnas.44.7.633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. CANELLAKIS E. S. Pyrimidine metabolism. II. Enzymatic pathways of uracil anabolism. J Biol Chem. 1957 Jul;227(1):329–338. [PubMed] [Google Scholar]
  10. De Lucia P., Cairns J. Isolation of an E. coli strain with a mutation affecting DNA polymerase. Nature. 1969 Dec 20;224(5225):1164–1166. doi: 10.1038/2241164a0. [DOI] [PubMed] [Google Scholar]
  11. FLAKS J. G., COHEN S. S. The enzymic synthesis of 5-hydroxymethyldeoxycytidylic acid. Biochim Biophys Acta. 1957 Sep;25(3):667–668. doi: 10.1016/0006-3002(57)90553-x. [DOI] [PubMed] [Google Scholar]
  12. FLAKS J. G., ERWIN M. J., BUCHANAN J. M. Biosynthesis of the purines. XVI. The synthesis of adenosine 5'-phosphate and 5-amino-4-imidazolecarboxamide ribotide by a nucleotide pyrophosphorylase. J Biol Chem. 1957 Sep;228(1):201–213. [PubMed] [Google Scholar]
  13. Frederiksen S. Effect of 2-aminopurine and 2-aminopurine 2'-deoxyriboside on nucleic acid synthesis in Ehrlich ascites cells in vitro. Biochem Pharmacol. 1965 May;14(5):651–660. doi: 10.1016/0006-2952(65)90083-3. [DOI] [PubMed] [Google Scholar]
  14. GINSBURG A. A deoxyribokinase from Lactobacillus plantarum. J Biol Chem. 1959 Mar;234(3):481–487. [PubMed] [Google Scholar]
  15. Hall Z. W., Lehman I. R. An in vitro transversion by a mutationally altered T4-induced DNA polymerase. J Mol Biol. 1968 Sep 28;36(3):321–333. doi: 10.1016/0022-2836(68)90158-7. [DOI] [PubMed] [Google Scholar]
  16. KALLE G. P., GOTS J. S. GENETIC ALTERATION OF ADENYLIC PYROPHOSPHORYLASE IN SALMONELLA. Science. 1963 Nov 8;142(3593):680–681. doi: 10.1126/science.142.3593.680. [DOI] [PubMed] [Google Scholar]
  17. KOCH A. L. Some enzymes of nucleoside metabolism of Escherichia coli. J Biol Chem. 1956 Nov;223(1):535–549. [PubMed] [Google Scholar]
  18. KORNBERG A., LIEBERMAN I., SIMMS E. S. Enzymatic synthesis of purine nucleotides. J Biol Chem. 1955 Jul;215(1):417–427. [PubMed] [Google Scholar]
  19. KORNBERG A., PRICER W. E., Jr Enzymatic phosphorylation of adenosine and 2,6-diaminopurine riboside. J Biol Chem. 1951 Dec;193(2):481–495. [PubMed] [Google Scholar]
  20. Krenitsky T. A., Neil S. M., Elion G. B., Hitchings G. H. Adenine phosphoribosyltransferase from monkey liver. Specificity and properties. J Biol Chem. 1969 Sep 10;244(17):4779–4784. [PubMed] [Google Scholar]
  21. Krenitsky T. A., Papaioannou R., Elion G. B. Human hypoxanthine phosphoribosyltransferase. I. Purification, properties, and specificity. J Biol Chem. 1969 Mar 10;244(5):1263–1270. [PubMed] [Google Scholar]
  22. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  23. Lindberg B., Klenow H., Hansen K. Some properties of partially purified mammalian adenosine kinase. J Biol Chem. 1967 Feb 10;242(3):350–356. [PubMed] [Google Scholar]
  24. MACNUTT W. S. The enzymically catalysed transfer of the deoxyribosyl group from one purine or pyrimidine to another. Biochem J. 1952 Jan;50(3):384–397. doi: 10.1042/bj0500384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Miller R. L., Bieber A. L. Substrate binding specificity and properties of inosine monophosphate: pyrophosphate phosphoribosyltransferase (EC 2.4.2.8) from Brewers yeast. Biochemistry. 1969 Feb;8(2):603–608. doi: 10.1021/bi00830a021. [DOI] [PubMed] [Google Scholar]
  26. NIRENBERG M. W., MATTHAEI J. H. The dependence of cell-free protein synthesis in E. coli upon naturally occurring or synthetic polyribonucleotides. Proc Natl Acad Sci U S A. 1961 Oct 15;47:1588–1602. doi: 10.1073/pnas.47.10.1588. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. OKAZAKI R., KORNBERG A. DEOXYTHYMIDINE KINASE OF ESCHERICHIA COLI. I. PURIFICATION AND SOME PROPERTIES OF THE ENZYME. J Biol Chem. 1964 Jan;239:269–274. [PubMed] [Google Scholar]
  28. OTT J. L., WERKMAN C. H. Coupled nucleoside phosphorylase reactions in Escherichia coli. Arch Biochem Biophys. 1957 Jul;69:264–276. doi: 10.1016/0003-9861(57)90491-5. [DOI] [PubMed] [Google Scholar]
  29. Oeschger M. P., Bessman M. J. Purification and properties of guanylate kinase from Escherichia coli. J Biol Chem. 1966 Nov 25;241(22):5452–5460. [PubMed] [Google Scholar]
  30. Pinto B., Touster O. Separation and modification of the phosphorolytic and ribosyl transfer activities of the purine nucleoside phosphorylase of Ehrlich ascites tumor cells. J Biol Chem. 1966 Feb 10;241(3):772–773. [PubMed] [Google Scholar]
  31. RICHARDSON C. C., SCHILDKRAUT C. L., APOSHIAN H. V., KORNBERG A. ENZYMATIC SYNTHESIS OF DEOXYRIBONUCLEIC ACID. XIV. FURTHER PURIFICATION AND PROPERTIES OF DEOXYRIBONUCLEIC ACID POLYMERASE OF ESCHERICHIA COLI. J Biol Chem. 1964 Jan;239:222–232. [PubMed] [Google Scholar]
  32. RUDNER R. Mutation as an error in base pairing. I. The mutagenicity of base analogues and their incorporation into the DNA of Salmonella typhimurium. Z Vererbungsl. 1961;92:336–360. [PubMed] [Google Scholar]
  33. SABLE H. Z. Phosphorylation of ribose and adenosine in yeast extracts. Proc Soc Exp Biol Med. 1950 Oct;75(1):215–219. doi: 10.3181/00379727-75-18149. [DOI] [PubMed] [Google Scholar]
  34. SULKOWSKI E., BJORK W., LASKOWSKI M., Sr A specific and nonspecific alkaline monophosphatase in the venom of Bothrops atrox and their occurrence in the purified venom phosphodiesterase. J Biol Chem. 1963 Jul;238:2477–2486. [PubMed] [Google Scholar]
  35. Speyer J. F. Mutagenic DNA polymerase. Biochem Biophys Res Commun. 1965 Oct 8;21(1):6–8. doi: 10.1016/0006-291x(65)90417-1. [DOI] [PubMed] [Google Scholar]
  36. Steuart C. D., Anand S. R., Bessman M. J. Studies on the synthesis of deoxyribonucleic acid. I. Further purification and properties of the deoxyribonucleic acid polymerase induced by infection of Escherichia coli with bacteriophage T5. J Biol Chem. 1968 Oct 25;243(20):5308–5318. [PubMed] [Google Scholar]
  37. WARAVDEKAR V. S., SASLAW L. D. A sensitive colorimetric method for the estimation of 2-deoxy sugars with the use of the malonaldehyde-thiobarbituric acid reaction. J Biol Chem. 1959 Aug;234(8):1945–1950. [PubMed] [Google Scholar]
  38. WHITFELD P. R. A method for the determination of nucleotide sequence in polyribonucleotides. Biochem J. 1954 Nov;58(3):390–396. doi: 10.1042/bj0580390. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Wacker A., Lodemann E., Gauri K., Chandra P. Synthesis and coding properties of 2-aminopurine polyribonucleotide. J Mol Biol. 1966 Jul;18(2):382–383. doi: 10.1016/s0022-2836(66)80255-3. [DOI] [PubMed] [Google Scholar]
  40. Ward D. C., Reich E., Stryer L. Fluorescence studies of nucleotides and polynucleotides. I. Formycin, 2-aminopurine riboside, 2,6-diaminopurine riboside, and their derivatives. J Biol Chem. 1969 Mar 10;244(5):1228–1237. [PubMed] [Google Scholar]

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