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. 1983 Mar 25;11(6):1773–1790. doi: 10.1093/nar/11.6.1773

Cloning of the gene topA encoding for DNA topoisomerase I and the physical mapping of the cysB-topA-trp region of Escherichia coli.

J C Wang, K Becherer
PMCID: PMC325835  PMID: 6300778

Abstract

The gene topA of Escherichia coli that encodes for DNA topoisomerase I has been cloned by a combination of genetic and radioimmunal screening. The gene has been mapped to be within a 3.4 Kb segment of the bacterial genome. The intracellular level of the enzyme in strains harboring extrachromosomal copies of topA gene increases with increasing copy number of the gene and the introduction of extrachromosomal copies of the topA gene truncated at its 3' side into a topA strain of E. coli does not significantly influence the expression of the chromosomal copy of topA. These results suggest that the expression of topA is not tightly regulated. Strains in which DNA topoisomerase I is overproduced grow significantly slower in broth and give smaller size colonies on agar plates. Physical mapping of a 20 Kb region containing cysB; topA and trp has also been carried out with a number of restriction enzymes; topA is found to be immediately adjacent to cysB and is separated from trp by a 7 Kb segment where no known gene resides.

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Selected References

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

  1. Barak Z., Gilvarg C. Triornithine-resistant strains of Escherichia coli. Isolation, definition, and genetic studies. J Biol Chem. 1974 Jan 10;249(1):143–148. [PubMed] [Google Scholar]
  2. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  3. Bouché J. P. Physical map of a 470 x 10(3) base-pair region flanking the terminus of DNA replication in the Escherichia coli K12 genome. J Mol Biol. 1982 Jan 5;154(1):1–20. doi: 10.1016/0022-2836(82)90413-2. [DOI] [PubMed] [Google Scholar]
  4. Broome S., Gilbert W. Immunological screening method to detect specific translation products. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2746–2749. doi: 10.1073/pnas.75.6.2746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brown P. O., Cozzarelli N. R. Catenation and knotting of duplex DNA by type 1 topoisomerases: a mechanistic parallel with type 2 topoisomerases. Proc Natl Acad Sci U S A. 1981 Feb;78(2):843–847. doi: 10.1073/pnas.78.2.843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Champoux J. J. DNA is linked to the rat liver DNA nicking-closing enzyme by a phosphodiester bond to tyrosine. J Biol Chem. 1981 May 25;256(10):4805–4809. [PubMed] [Google Scholar]
  7. Champoux J. J. Strand breakage by the DNA untwisting enzyme results in covalent attachment of the enzyme to DNA. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3800–3804. doi: 10.1073/pnas.74.9.3800. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Christie G. E., Platt T. Gene structure in the tryptophan operon of Escherichia coli. Nucleotide sequence of trpC and the flanking intercistronic regions. J Mol Biol. 1980 Oct 5;142(4):519–530. doi: 10.1016/0022-2836(80)90261-2. [DOI] [PubMed] [Google Scholar]
  9. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  10. Cozzarelli N. R. DNA gyrase and the supercoiling of DNA. Science. 1980 Feb 29;207(4434):953–960. doi: 10.1126/science.6243420. [DOI] [PubMed] [Google Scholar]
  11. Crawford I. P., Nichols B. P., Yanofsky C. Nucleotide sequence of the trpB gene in Escherichia coli and Salmonella typhimurium. J Mol Biol. 1980 Oct 5;142(4):489–502. doi: 10.1016/0022-2836(80)90259-4. [DOI] [PubMed] [Google Scholar]
  12. De Felice M., Guardiola J., Lamberti A., Iaccarino M. Escherichia coli K-12 mutants altered in the transport systems for oligo- and dipeptides. J Bacteriol. 1973 Nov;116(2):751–756. doi: 10.1128/jb.116.2.751-756.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Depew R. E., Liu L. F., Wang J. C. Interaction between DNA and Escherichia coli protein omega. Formation of a complex between single-stranded DNA and omega protein. J Biol Chem. 1978 Jan 25;253(2):511–518. [PubMed] [Google Scholar]
  14. DiNardo S., Voelkel K. A., Sternglanz R., Reynolds A. E., Wright A. Escherichia coli DNA topoisomerase I mutants have compensatory mutations in DNA gyrase genes. Cell. 1982 Nov;31(1):43–51. doi: 10.1016/0092-8674(82)90403-2. [DOI] [PubMed] [Google Scholar]
  15. Dubnau E., Lenny A. B., Margolin P. Nonsense mutations of the supX locus: further characterization of the supX mutant phenotype. Mol Gen Genet. 1973 Nov 12;126(3):191–200. doi: 10.1007/BF00267530. [DOI] [PubMed] [Google Scholar]
  16. Dubnau E., Margolin P. Suppression of promoter mutations by the pleiotropic supx mutations. Mol Gen Genet. 1972;117(2):91–112. doi: 10.1007/BF00267607. [DOI] [PubMed] [Google Scholar]
  17. Gellert M. DNA topoisomerases. Annu Rev Biochem. 1981;50:879–910. doi: 10.1146/annurev.bi.50.070181.004311. [DOI] [PubMed] [Google Scholar]
  18. Gellert M., Mizuuchi K., O'Dea M. H., Itoh T., Tomizawa J. I. Nalidixic acid resistance: a second genetic character involved in DNA gyrase activity. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4772–4776. doi: 10.1073/pnas.74.11.4772. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gray H. B., Jr, Ostrander D. A., Hodnett J. L., Legerski R. J., Robberson D. L. Extracellular nucleases of Pseudomonas BAL 31. I. Characterization of single strand-specific deoxyriboendonuclease and double-strand deoxyriboexonuclease activities. Nucleic Acids Res. 1975 Sep;2(9):1459–1492. doi: 10.1093/nar/2.9.1459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Guerry P., LeBlanc D. J., Falkow S. General method for the isolation of plasmid deoxyribonucleic acid. J Bacteriol. 1973 Nov;116(2):1064–1066. doi: 10.1128/jb.116.2.1064-1066.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hickson F. T., Roth T. F., Helinski D. R. Circular DNA forms of a bacterial sex factor. Proc Natl Acad Sci U S A. 1967 Oct;58(4):1731–1738. doi: 10.1073/pnas.58.4.1731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  23. Hsieh T., Wang J. C. Physicochomecial studies on interactions between DNA and RNA polymerase. Isolation and mapping of a T7 DNA fragment containing the early promoters for Escherichia coli RNA polymerase. Biochemistry. 1976 Dec 28;15(26):5776–5783. doi: 10.1021/bi00671a014. [DOI] [PubMed] [Google Scholar]
  24. Humphreys G. O., Willshaw G. A., Anderson E. S. A simple method for the preparation of large quantities of pure plasmid DNA. Biochim Biophys Acta. 1975 Apr 2;383(4):457–463. doi: 10.1016/0005-2787(75)90318-4. [DOI] [PubMed] [Google Scholar]
  25. Jovin T. M., Englund P. T., Kornberg A. Enzymatic synthesis of deoxyribonucleic acid. XXVII. Chemical modifications of deoxyribonucleic acid polymerase. J Biol Chem. 1969 Jun 10;244(11):3009–3018. [PubMed] [Google Scholar]
  26. Kahn M., Kolter R., Thomas C., Figurski D., Meyer R., Remaut E., Helinski D. R. Plasmid cloning vehicles derived from plasmids ColE1, F, R6K, and RK2. Methods Enzymol. 1979;68:268–280. doi: 10.1016/0076-6879(79)68019-9. [DOI] [PubMed] [Google Scholar]
  27. Kirkegaard K., Wang J. C. Escherichia coli DNA topoisomerase I catalyzed linking of single-stranded rings of complementary base sequences. Nucleic Acids Res. 1978 Oct;5(10):3811–3820. doi: 10.1093/nar/5.10.3811. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Lenny A. B., Margolin P. Locations of the opp and supX genes of Salmonella typhimurium and Escherichia coli. J Bacteriol. 1980 Aug;143(2):747–752. doi: 10.1128/jb.143.2.747-752.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Meyer T. F., Mlawer N., So M. Pilus expression in Neisseria gonorrhoeae involves chromosomal rearrangement. Cell. 1982 Aug;30(1):45–52. doi: 10.1016/0092-8674(82)90010-1. [DOI] [PubMed] [Google Scholar]
  30. Mukai F. H., Margolin P. ANALYSIS OF UNLINKED SUPPRESSORS OF AN O degrees MUTATION IN SALMONELLA. Proc Natl Acad Sci U S A. 1963 Jul;50(1):140–148. doi: 10.1073/pnas.50.1.140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nichols B. P., Miozzari G. F., van Cleemput M., Bennett G. N., Yanofsky C. Nucleotide sequences of the trpG regions of Escherichia coli, Shigella dysenteriae, Salmonella typhimurium and Serratia marcescens. J Mol Biol. 1980 Oct 5;142(4):503–517. doi: 10.1016/0022-2836(80)90260-0. [DOI] [PubMed] [Google Scholar]
  32. Nichols B. P., Yanofsky C. Nucleotide sequences of trpA of Salmonella typhimurium and Escherichia coli: an evolutionary comparison. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5244–5248. doi: 10.1073/pnas.76.10.5244. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Nichols B. P., van Cleemput M., Yanofsky C. Nucleotide sequence of Escherichia coli trpE. Anthranilate synthetase component I contains no tryptophan residues. J Mol Biol. 1981 Feb 15;146(1):45–54. doi: 10.1016/0022-2836(81)90365-x. [DOI] [PubMed] [Google Scholar]
  34. Paoletti C., LePecq J. B., Lehman I. R. The use of ethidium bromide-circular DNA complexes for the fluorometric analysis of breakage and joining of DNA. J Mol Biol. 1971 Jan 14;55(1):75–100. doi: 10.1016/0022-2836(71)90282-8. [DOI] [PubMed] [Google Scholar]
  35. Pruss G. J., Manes S. H., Drlica K. Escherichia coli DNA topoisomerase I mutants: increased supercoiling is corrected by mutations near gyrase genes. Cell. 1982 Nov;31(1):35–42. doi: 10.1016/0092-8674(82)90402-0. [DOI] [PubMed] [Google Scholar]
  36. Pulleyblank D. E., Shure M., Tang D., Vinograd J., Vosberg H. P. Action of nicking-closing enzyme on supercoiled and nonsupercoiled closed circular DNA: formation of a Boltzmann distribution of topological isomers. Proc Natl Acad Sci U S A. 1975 Nov;72(11):4280–4284. doi: 10.1073/pnas.72.11.4280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Roberts T. M., Kacich R., Ptashne M. A general method for maximizing the expression of a cloned gene. Proc Natl Acad Sci U S A. 1979 Feb;76(2):760–764. doi: 10.1073/pnas.76.2.760. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Smith H. O., Birnstiel M. L. A simple method for DNA restriction site mapping. Nucleic Acids Res. 1976 Sep;3(9):2387–2398. doi: 10.1093/nar/3.9.2387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  40. Sternglanz R., DiNardo S., Voelkel K. A., Nishimura Y., Hirota Y., Becherer K., Zumstein L., Wang J. C. Mutations in the gene coding for Escherichia coli DNA topoisomerase I affect transcription and transposition. Proc Natl Acad Sci U S A. 1981 May;78(5):2747–2751. doi: 10.1073/pnas.78.5.2747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sugino A., Peebles C. L., Kreuzer K. N., Cozzarelli N. R. Mechanism of action of nalidixic acid: purification of Escherichia coli nalA gene product and its relationship to DNA gyrase and a novel nicking-closing enzyme. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4767–4771. doi: 10.1073/pnas.74.11.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sutcliffe J. G. Complete nucleotide sequence of the Escherichia coli plasmid pBR322. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):77–90. doi: 10.1101/sqb.1979.043.01.013. [DOI] [PubMed] [Google Scholar]
  43. Trucksis M., Depew R. E. Identification and localization of a gene that specifies production of Escherichia coli DNA topoisomerase I. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2164–2168. doi: 10.1073/pnas.78.4.2164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Trucksis M., Golub E. I., Zabel D. J., Depew R. E. Escherichia coli and Salmonella typhimurium supX genes specify deoxyribonucleic acid topoisomerase I. J Bacteriol. 1981 Aug;147(2):679–681. doi: 10.1128/jb.147.2.679-681.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tse Y. C., Kirkegaard K., Wang J. C. Covalent bonds between protein and DNA. Formation of phosphotyrosine linkage between certain DNA topoisomerases and DNA. J Biol Chem. 1980 Jun 25;255(12):5560–5565. [PubMed] [Google Scholar]
  46. Tse Y., Wang J. C. E. coli and M. luteus DNA topoisomerase I can catalyze catenation of decatenation of double-stranded DNA rings. Cell. 1980 Nov;22(1 Pt 1):269–276. doi: 10.1016/0092-8674(80)90174-9. [DOI] [PubMed] [Google Scholar]
  47. Wang J. C. Interaction between DNA and an Escherichia coli protein omega. J Mol Biol. 1971 Feb 14;55(3):523–533. doi: 10.1016/0022-2836(71)90334-2. [DOI] [PubMed] [Google Scholar]
  48. Wickner W., Brutlag D., Schekman R., Kornberg A. RNA synthesis initiates in vitro conversion of M13 DNA to its replicative form. Proc Natl Acad Sci U S A. 1972 Apr;69(4):965–969. doi: 10.1073/pnas.69.4.965. [DOI] [PMC free article] [PubMed] [Google Scholar]

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