Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Dec 15;89(24):12108–12111. doi: 10.1073/pnas.89.24.12108

Activity of the Hsp70 chaperone complex--DnaK, DnaJ, and GrpE--in initiating phage lambda DNA replication by sequestering and releasing lambda P protein.

H J Hoffmann 1, S K Lyman 1, C Lu 1, M A Petit 1, H Echols 1
PMCID: PMC50707  PMID: 1361234

Abstract

Initiation of DNA replication by phage lambda requires the ordered assembly and disassembly of a specialized nucleoprotein structure at the origin of replication. In the disassembly pathway, a set of Escherichia coli heat shock proteins termed the Hsp70 complex--DnaK, DnaJ, and GrpE--act with ATP to release lambda P protein from the nucleo-protein complex, freeing the DnaB helicase for its DNA-unwinding reaction. To investigate the mechanism of the release reaction, we have examined the interaction between P and the three heat shock proteins by glycerol gradient sedimentation and gel electrophoresis. We have discovered an ATP-dependent ternary interaction between P, DnaK, and DnaJ; this P.DnaK.DnaJ complex is dissociated by GrpE. We have concluded that the function of the Hsp70 complex in sequestering and releasing P protein provides for the critical step in the disassembly pathway. Based on our data and other work on protein folding, the formation of the P.DnaK.DnaJ complex might involve a conformational shift to a folding intermediate of P.

Full text

PDF
12108

Images in this article

12109Image
on p.12109
12110Image
on p.12110
12110Image
on p.12110

Selected References

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

  1. Alfano C., McMacken R. Heat shock protein-mediated disassembly of nucleoprotein structures is required for the initiation of bacteriophage lambda DNA replication. J Biol Chem. 1989 Jun 25;264(18):10709–10718. [PubMed] [Google Scholar]
  2. Dodson M., Echols H., Wickner S., Alfano C., Mensa-Wilmot K., Gomes B., LeBowitz J., Roberts J. D., McMacken R. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda: localized unwinding of duplex DNA by a six-protein reaction. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7638–7642. doi: 10.1073/pnas.83.20.7638. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dodson M., McMacken R., Echols H. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda. Protein association and disassociation reactions responsible for localized initiation of replication. J Biol Chem. 1989 Jun 25;264(18):10719–10725. [PubMed] [Google Scholar]
  4. Dodson M., Roberts J., McMacken R., Echols H. Specialized nucleoprotein structures at the origin of replication of bacteriophage lambda: complexes with lambda O protein and with lambda O, lambda P, and Escherichia coli DnaB proteins. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4678–4682. doi: 10.1073/pnas.82.14.4678. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Georgopoulos C. The emergence of the chaperone machines. Trends Biochem Sci. 1992 Aug;17(8):295–299. doi: 10.1016/0968-0004(92)90439-g. [DOI] [PubMed] [Google Scholar]
  7. Gething M. J., Sambrook J. Protein folding in the cell. Nature. 1992 Jan 2;355(6355):33–45. doi: 10.1038/355033a0. [DOI] [PubMed] [Google Scholar]
  8. Johnson C., Chandrasekhar G. N., Georgopoulos C. Escherichia coli DnaK and GrpE heat shock proteins interact both in vivo and in vitro. J Bacteriol. 1989 Mar;171(3):1590–1596. doi: 10.1128/jb.171.3.1590-1596.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Langer T., Lu C., Echols H., Flanagan J., Hayer M. K., Hartl F. U. Successive action of DnaK, DnaJ and GroEL along the pathway of chaperone-mediated protein folding. Nature. 1992 Apr 23;356(6371):683–689. doi: 10.1038/356683a0. [DOI] [PubMed] [Google Scholar]
  10. Liberek K., Georgopoulos C., Zylicz M. Role of the Escherichia coli DnaK and DnaJ heat shock proteins in the initiation of bacteriophage lambda DNA replication. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6632–6636. doi: 10.1073/pnas.85.18.6632. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Liberek K., Marszalek J., Ang D., Georgopoulos C., Zylicz M. Escherichia coli DnaJ and GrpE heat shock proteins jointly stimulate ATPase activity of DnaK. Proc Natl Acad Sci U S A. 1991 Apr 1;88(7):2874–2878. doi: 10.1073/pnas.88.7.2874. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Liberek K., Osipiuk J., Zylicz M., Ang D., Skorko J., Georgopoulos C. Physical interactions between bacteriophage and Escherichia coli proteins required for initiation of lambda DNA replication. J Biol Chem. 1990 Feb 25;265(6):3022–3029. [PubMed] [Google Scholar]
  13. Mallory J. B., Alfano C., McMacken R. Host virus interactions in the initiation of bacteriophage lambda DNA replication. Recruitment of Escherichia coli DnaB helicase by lambda P replication protein. J Biol Chem. 1990 Aug 5;265(22):13297–13307. [PubMed] [Google Scholar]
  14. Marians K. J. Prokaryotic DNA replication. Annu Rev Biochem. 1992;61:673–719. doi: 10.1146/annurev.bi.61.070192.003325. [DOI] [PubMed] [Google Scholar]
  15. Mensa-Wilmot K., Seaby R., Alfano C., Wold M. C., Gomes B., McMacken R. Reconstitution of a nine-protein system that initiates bacteriophage lambda DNA replication. J Biol Chem. 1989 Feb 15;264(5):2853–2861. [PubMed] [Google Scholar]
  16. Osipiuk J., Zylicz M. Role of the Escherichia coli grpE heat shock protein in the initiation of bacteriophage lambda DNA replication. Acta Biochim Pol. 1991;38(1):191–200. [PubMed] [Google Scholar]
  17. Roberts J. D., McMacken R. The bacteriophage lambda O replication protein: isolation and characterization of the amplified initiator. Nucleic Acids Res. 1983 Nov 11;11(21):7435–7452. [PMC free article] [PubMed] [Google Scholar]
  18. Rothman J. E. Polypeptide chain binding proteins: catalysts of protein folding and related processes in cells. Cell. 1989 Nov 17;59(4):591–601. doi: 10.1016/0092-8674(89)90005-6. [DOI] [PubMed] [Google Scholar]
  19. Saito H., Uchida H. Initiation of the DNA replication of bacteriophage lambda in Escherichia coli K12. J Mol Biol. 1977 Jun 15;113(1):1–25. doi: 10.1016/0022-2836(77)90038-9. [DOI] [PubMed] [Google Scholar]
  20. Straus D., Walter W., Gross C. A. DnaK, DnaJ, and GrpE heat shock proteins negatively regulate heat shock gene expression by controlling the synthesis and stability of sigma 32. Genes Dev. 1990 Dec;4(12A):2202–2209. doi: 10.1101/gad.4.12a.2202. [DOI] [PubMed] [Google Scholar]
  21. Tsurimoto T., Matsubara K. Purified bacteriophage lambda O protein binds to four repeating sequences at the lambda replication origin. Nucleic Acids Res. 1981 Apr 24;9(8):1789–1799. doi: 10.1093/nar/9.8.1789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wickner S. H. DNA replication proteins of Escherichia coli and phage lambda. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 1):303–310. doi: 10.1101/sqb.1979.043.01.037. [DOI] [PubMed] [Google Scholar]
  23. Wickner S. H., Zahn K. Characterization of the DNA binding domain of bacteriophage lambda O protein. J Biol Chem. 1986 Jun 5;261(16):7537–7543. [PubMed] [Google Scholar]
  24. Wickner S., Hoskins J., McKenney K. Monomerization of RepA dimers by heat shock proteins activates binding to DNA replication origin. Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):7903–7907. doi: 10.1073/pnas.88.18.7903. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Zahn K., Blattner F. R. Direct evidence for DNA bending at the lambda replication origin. Science. 1987 Apr 24;236(4800):416–422. doi: 10.1126/science.2951850. [DOI] [PubMed] [Google Scholar]
  26. Zylicz M., Ang D., Georgopoulos C. The grpE protein of Escherichia coli. Purification and properties. J Biol Chem. 1987 Dec 25;262(36):17437–17442. [PubMed] [Google Scholar]
  27. Zylicz M., Ang D., Liberek K., Georgopoulos C. Initiation of lambda DNA replication with purified host- and bacteriophage-encoded proteins: the role of the dnaK, dnaJ and grpE heat shock proteins. EMBO J. 1989 May;8(5):1601–1608. doi: 10.1002/j.1460-2075.1989.tb03544.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Zylicz M., Georgopoulos C. Purification and properties of the Escherichia coli dnaK replication protein. J Biol Chem. 1984 Jul 25;259(14):8820–8825. [PubMed] [Google Scholar]
  29. Zylicz M., Yamamoto T., McKittrick N., Sell S., Georgopoulos C. Purification and properties of the dnaJ replication protein of Escherichia coli. J Biol Chem. 1985 Jun 25;260(12):7591–7598. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES