Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1993 Dec;12(12):4547–4554. doi: 10.1002/j.1460-2075.1993.tb06143.x

Open-complex formation by the host initiator, DnaA, at the origin of P1 plasmid replication.

G Mukhopadhyay 1, K M Carr 1, J M Kaguni 1, D K Chattoraj 1
PMCID: PMC413885  PMID: 8223464

Abstract

Replication of P1 plasmid requires both the plasmid-specific initiator, RepA, and the host initiator, DnaA. Here we show that DnaA can make the P1 origin reactive to the single-strand specific reagents KMnO4 and mung bean nuclease. Addition of RepA further increased the KMnO4 reactivity of the origin, although RepA alone did not influence the reaction. The increased reactivity implies that the two initiators interact in some way to alter the origin conformation. The KMnO4 reactivity was restricted to one strand of the origin. We suggest that the roles of DnaA in P1 plasmid and bacterial replication are similar: origin opening and loading of the DnaB helicase. The strand-bias in chemical reactivity at the P1 origin most likely indicates that only one of the strands is used for the loading of DnaB, a scenario consistent with the unidirectional replication of the plasmid.

Full text

PDF
4547

Images in this article

4548Image
on p.4548
4549Image
on p.4549
4549Image
on p.4549
4550Image
on p.4550
4551Image
on p.4551
4552Image
on p.4552
4553Image
on p.4553

Selected References

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

  1. Abeles A. L. P1 plasmid replication. Purification and DNA-binding activity of the replication protein RepA. J Biol Chem. 1986 Mar 15;261(8):3548–3555. [PubMed] [Google Scholar]
  2. Abeles A. L., Reaves L. D., Austin S. J. A single DnaA box is sufficient for initiation from the P1 plasmid origin. J Bacteriol. 1990 Aug;172(8):4386–4391. doi: 10.1128/jb.172.8.4386-4391.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Abeles A. L., Snyder K. M., Chattoraj D. K. P1 plasmid replication: replicon structure. J Mol Biol. 1984 Mar 5;173(3):307–324. doi: 10.1016/0022-2836(84)90123-2. [DOI] [PubMed] [Google Scholar]
  4. Austin S. J., Eichorn B. G. Random diffusion can account for topA-dependent suppression of partition defects in low-copy-number plasmids. J Bacteriol. 1992 Aug;174(16):5190–5195. doi: 10.1128/jb.174.16.5190-5195.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Borowiec J. A., Zhang L., Sasse-Dwight S., Gralla J. D. DNA supercoiling promotes formation of a bent repression loop in lac DNA. J Mol Biol. 1987 Jul 5;196(1):101–111. doi: 10.1016/0022-2836(87)90513-4. [DOI] [PubMed] [Google Scholar]
  6. Bramhill D., Kornberg A. A model for initiation at origins of DNA replication. Cell. 1988 Sep 23;54(7):915–918. doi: 10.1016/0092-8674(88)90102-x. [DOI] [PubMed] [Google Scholar]
  7. Bramhill D., Kornberg A. Duplex opening by dnaA protein at novel sequences in initiation of replication at the origin of the E. coli chromosome. Cell. 1988 Mar 11;52(5):743–755. doi: 10.1016/0092-8674(88)90412-6. [DOI] [PubMed] [Google Scholar]
  8. DasGupta S., Mukhopadhyay G., Papp P. P., Lewis M. S., Chattoraj D. K. Activation of DNA binding by the monomeric form of the P1 replication initiator RepA by heat shock proteins DnaJ and DnaK. J Mol Biol. 1993 Jul 5;232(1):23–34. doi: 10.1006/jmbi.1993.1367. [DOI] [PubMed] [Google Scholar]
  9. Echols H. Nucleoprotein structures initiating DNA replication, transcription, and site-specific recombination. J Biol Chem. 1990 Sep 5;265(25):14697–14700. [PubMed] [Google Scholar]
  10. Fuller R. S., Funnell B. E., Kornberg A. The dnaA protein complex with the E. coli chromosomal replication origin (oriC) and other DNA sites. Cell. 1984 Oct;38(3):889–900. doi: 10.1016/0092-8674(84)90284-8. [DOI] [PubMed] [Google Scholar]
  11. Georgopoulos C. The E. coli dnaA initiation protein: a protein for all seasons. Trends Genet. 1989 Oct;5(10):319–321. doi: 10.1016/0168-9525(89)90118-2. [DOI] [PubMed] [Google Scholar]
  12. Gille H., Messer W. Localized DNA melting and structural pertubations in the origin of replication, oriC, of Escherichia coli in vitro and in vivo. EMBO J. 1991 Jun;10(6):1579–1584. doi: 10.1002/j.1460-2075.1991.tb07678.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hansen E. B., Yarmolinsky M. B. Host participation in plasmid maintenance: dependence upon dnaA of replicons derived from P1 and F. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4423–4427. doi: 10.1073/pnas.83.12.4423. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Holz A., Schaefer C., Gille H., Jueterbock W. R., Messer W. Mutations in the DnaA binding sites of the replication origin of Escherichia coli. Mol Gen Genet. 1992 May;233(1-2):81–88. doi: 10.1007/BF00587564. [DOI] [PubMed] [Google Scholar]
  15. Hwang D. S., Crooke E., Kornberg A. Aggregated dnaA protein is dissociated and activated for DNA replication by phospholipase or dnaK protein. J Biol Chem. 1990 Nov 5;265(31):19244–19248. [PubMed] [Google Scholar]
  16. Hwang D. S., Kaguni J. M. Purification and characterization of the dnaA46 gene product. J Biol Chem. 1988 Aug 5;263(22):10625–10632. [PubMed] [Google Scholar]
  17. Hwang D. S., Kornberg A. Opposed actions of regulatory proteins, DnaA and IciA, in opening the replication origin of Escherichia coli. J Biol Chem. 1992 Nov 15;267(32):23087–23091. [PubMed] [Google Scholar]
  18. Itoh Y., Terawaki Y. Replication properties of mini-Rts1 derivatives deleted for DnaA boxes in the replication origin. Plasmid. 1989 May;21(3):242–246. doi: 10.1016/0147-619x(89)90048-6. [DOI] [PubMed] [Google Scholar]
  19. Jensen M. R., Løbner-Olesen A., Rasmussen K. V. Escherichia coli minichromosomes: random segregation and absence of copy number control. J Mol Biol. 1990 Sep 20;215(2):257–265. doi: 10.1016/S0022-2836(05)80344-4. [DOI] [PubMed] [Google Scholar]
  20. Kainz M., Roberts J. Structure of transcription elongation complexes in vivo. Science. 1992 Feb 14;255(5046):838–841. doi: 10.1126/science.1536008. [DOI] [PubMed] [Google Scholar]
  21. Kowalski D., Eddy M. J. The DNA unwinding element: a novel, cis-acting component that facilitates opening of the Escherichia coli replication origin. EMBO J. 1989 Dec 20;8(13):4335–4344. doi: 10.1002/j.1460-2075.1989.tb08620.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Løbner-Olesen A., Skarstad K., Hansen F. G., von Meyenburg K., Boye E. The DnaA protein determines the initiation mass of Escherichia coli K-12. Cell. 1989 Jun 2;57(5):881–889. doi: 10.1016/0092-8674(89)90802-7. [DOI] [PubMed] [Google Scholar]
  23. Masai H., Arai K. RepA and DnaA proteins are required for initiation of R1 plasmid replication in vitro and interact with the oriR sequence. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4781–4785. doi: 10.1073/pnas.84.14.4781. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Masai H., Nomura N., Arai K. The ABC-primosome. A novel priming system employing dnaA, dnaB, dnaC, and primase on a hairpin containing a dnaA box sequence. J Biol Chem. 1990 Sep 5;265(25):15134–15144. [PubMed] [Google Scholar]
  25. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mukhopadhyay G., Chattoraj D. K. Conformation of the origin of P1 plasmid replication. Initiator protein induced wrapping and intrinsic unstacking. J Mol Biol. 1993 May 5;231(1):19–28. doi: 10.1006/jmbi.1993.1253. [DOI] [PubMed] [Google Scholar]
  27. Ortega-Jiménez S., Giraldo-Suárez R., Fernández-Tresguerres M. E., Berzal-Herranz A., Díaz-Orejas R. DnaA dependent replication of plasmid R1 occurs in the presence of point mutations that disrupt the dnaA box of oriR. Nucleic Acids Res. 1992 May 25;20(10):2547–2551. doi: 10.1093/nar/20.10.2547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Pal S. K., Mason R. J., Chattoraj D. K. P1 plasmid replication. Role of initiator titration in copy number control. J Mol Biol. 1986 Nov 20;192(2):275–285. doi: 10.1016/0022-2836(86)90364-5. [DOI] [PubMed] [Google Scholar]
  29. Prentki P., Chandler M., Caro L. Replication of prophage P1 during the cell cycle of Escherichia coli. Mol Gen Genet. 1977 Mar 28;152(1):71–76. doi: 10.1007/BF00264942. [DOI] [PubMed] [Google Scholar]
  30. Sasse-Dwight S., Gralla J. D. KMnO4 as a probe for lac promoter DNA melting and mechanism in vivo. J Biol Chem. 1989 May 15;264(14):8074–8081. [PubMed] [Google Scholar]
  31. Seufert W., Dobrinski B., Lurz R., Messer W. Functionality of the dnaA protein binding site in DNA replication is orientation-dependent. J Biol Chem. 1988 Feb 25;263(6):2719–2723. [PubMed] [Google Scholar]
  32. Sozhamannan S., Chattoraj D. K. Heat shock proteins DnaJ, DnaK, and GrpE stimulate P1 plasmid replication by promoting initiator binding to the origin. J Bacteriol. 1993 Jun;175(11):3546–3555. doi: 10.1128/jb.175.11.3546-3555.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Stenzel T. T., MacAllister T., Bastia D. Cooperativity at a distance promoted by the combined action of two replication initiator proteins and a DNA bending protein at the replication origin of pSC101. Genes Dev. 1991 Aug;5(8):1453–1463. doi: 10.1101/gad.5.8.1453. [DOI] [PubMed] [Google Scholar]
  34. Suh W. C., Ross W., Record M. T., Jr Two open complexes and a requirement for Mg2+ to open the lambda PR transcription start site. Science. 1993 Jan 15;259(5093):358–361. doi: 10.1126/science.8420002. [DOI] [PubMed] [Google Scholar]
  35. Wickner S. H., Chattoraj D. K. Replication of mini-P1 plasmid DNA in vitro requires two initiation proteins, encoded by the repA gene of phage P1 and the dnaA gene of Escherichia coli. Proc Natl Acad Sci U S A. 1987 Jun;84(11):3668–3672. doi: 10.1073/pnas.84.11.3668. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wickner S., Hoskins J., Chattoraj D., McKenney K. Deletion analysis of the mini-P1 plasmid origin of replication and the role of Escherichia coli DnaA protein. J Biol Chem. 1990 Jul 15;265(20):11622–11627. [PubMed] [Google Scholar]
  37. Wickner S., Hoskins J., McKenney K. Function of DnaJ and DnaK as chaperones in origin-specific DNA binding by RepA. Nature. 1991 Mar 14;350(6314):165–167. doi: 10.1038/350165a0. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES