Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Jun;178(12):3608–3613. doi: 10.1128/jb.178.12.3608-3613.1996

Involvement of the DnaK-DnaJ-GrpE chaperone team in protein secretion in Escherichia coli.

J Wild 1, P Rossmeissl 1, W A Walter 1, C A Gross 1
PMCID: PMC178133  PMID: 8655561

Abstract

We used depletion studies designed to further investigate the role of the DnaK, DnaJ, and GrpE heat shock proteins in the SecB-dependent and SecB-independent secretion pathways. Our previous finding that SecB-deficient strains containing the grpE280 mutation were still secretion proficient raised the possibility that GrpE was not involved in this secretory pathway. Using depletion studies, we now demonstrate a requirement for GrpE in this pathway. In addition, depletion studies demonstrate that while DnaK, DnaJ, and GrpE are involved in the secretion of the SecB-independent proteins (alkaline phosphatase, ribose-binding protein, and beta-lactamase), they are not the primary chaperones in this process.

Full Text

The Full Text of this article is available as a PDF (505.8 KB).

Selected References

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

  1. Alfano C., McMacken R. Ordered assembly of nucleoprotein structures at the bacteriophage lambda replication origin during the initiation of DNA replication. J Biol Chem. 1989 Jun 25;264(18):10699–10708. [PubMed] [Google Scholar]
  2. Alix J. H., Guérin M. F. Mutant DnaK chaperones cause ribosome assembly defects in Escherichia coli. Proc Natl Acad Sci U S A. 1993 Oct 15;90(20):9725–9729. doi: 10.1073/pnas.90.20.9725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ang D., Georgopoulos C. The heat-shock-regulated grpE gene of Escherichia coli is required for bacterial growth at all temperatures but is dispensable in certain mutant backgrounds. J Bacteriol. 1989 May;171(5):2748–2755. doi: 10.1128/jb.171.5.2748-2755.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bieker-Brady K., Silhavy T. J. Suppressor analysis suggests a multistep, cyclic mechanism for protein secretion in Escherichia coli. EMBO J. 1992 Sep;11(9):3165–3174. doi: 10.1002/j.1460-2075.1992.tb05393.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bieker K. L., Phillips G. J., Silhavy T. J. The sec and prl genes of Escherichia coli. J Bioenerg Biomembr. 1990 Jun;22(3):291–310. doi: 10.1007/BF00763169. [DOI] [PubMed] [Google Scholar]
  6. Bolliger L., Deloche O., Glick B. S., Georgopoulos C., Jenö P., Kronidou N., Horst M., Morishima N., Schatz G. A mitochondrial homolog of bacterial GrpE interacts with mitochondrial hsp70 and is essential for viability. EMBO J. 1994 Apr 15;13(8):1998–2006. doi: 10.1002/j.1460-2075.1994.tb06469.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bukau B., Walker G. C. Delta dnaK52 mutants of Escherichia coli have defects in chromosome segregation and plasmid maintenance at normal growth temperatures. J Bacteriol. 1989 Nov;171(11):6030–6038. doi: 10.1128/jb.171.11.6030-6038.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Casadaban M. J., Cohen S. N. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. J Mol Biol. 1980 Apr;138(2):179–207. doi: 10.1016/0022-2836(80)90283-1. [DOI] [PubMed] [Google Scholar]
  9. Emr S. D., Hanley-Way S., Silhavy T. J. Suppressor mutations that restore export of a protein with a defective signal sequence. Cell. 1981 Jan;23(1):79–88. doi: 10.1016/0092-8674(81)90272-5. [DOI] [PubMed] [Google Scholar]
  10. Gamer J., Bujard H., Bukau B. Physical interaction between heat shock proteins DnaK, DnaJ, and GrpE and the bacterial heat shock transcription factor sigma 32. Cell. 1992 May 29;69(5):833–842. doi: 10.1016/0092-8674(92)90294-m. [DOI] [PubMed] [Google Scholar]
  11. Gannon P. M., Li P., Kumamoto C. A. The mature portion of Escherichia coli maltose-binding protein (MBP) determines the dependence of MBP on SecB for export. J Bacteriol. 1989 Feb;171(2):813–818. doi: 10.1128/jb.171.2.813-818.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gragerov A., Nudler E., Komissarova N., Gaitanaris G. A., Gottesman M. E., Nikiforov V. Cooperation of GroEL/GroES and DnaK/DnaJ heat shock proteins in preventing protein misfolding in Escherichia coli. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10341–10344. doi: 10.1073/pnas.89.21.10341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Grossman A. D., Straus D. B., Walter W. A., Gross C. A. Sigma 32 synthesis can regulate the synthesis of heat shock proteins in Escherichia coli. Genes Dev. 1987 Apr;1(2):179–184. doi: 10.1101/gad.1.2.179. [DOI] [PubMed] [Google Scholar]
  14. Hannavy K., Rospert S., Schatz G. Protein import into mitochondria: a paradigm for the translocation of polypeptides across membranes. Curr Opin Cell Biol. 1993 Aug;5(4):694–700. doi: 10.1016/0955-0674(93)90142-d. [DOI] [PubMed] [Google Scholar]
  15. Hendrick J. P., Hartl F. U. Molecular chaperone functions of heat-shock proteins. Annu Rev Biochem. 1993;62:349–384. doi: 10.1146/annurev.bi.62.070193.002025. [DOI] [PubMed] [Google Scholar]
  16. Hoffmann H. J., Lyman S. K., Lu C., Petit M. A., Echols H. Activity of the Hsp70 chaperone complex--DnaK, DnaJ, and GrpE--in initiating phage lambda DNA replication by sequestering and releasing lambda P protein. Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):12108–12111. doi: 10.1073/pnas.89.24.12108. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]
  18. Kandror O., Sherman M., Rhode M., Goldberg A. L. Trigger factor is involved in GroEL-dependent protein degradation in Escherichia coli and promotes binding of GroEL to unfolded proteins. EMBO J. 1995 Dec 1;14(23):6021–6027. doi: 10.1002/j.1460-2075.1995.tb00290.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kawasaki Y., Wada C., Yura T. Roles of Escherichia coli heat shock proteins DnaK, DnaJ and GrpE in mini-F plasmid replication. Mol Gen Genet. 1990 Jan;220(2):277–282. doi: 10.1007/BF00260494. [DOI] [PubMed] [Google Scholar]
  20. Keller J. A., Simon L. D. Divergent effects of a dnaK mutation on abnormal protein degradation in Escherichia coli. Mol Microbiol. 1988 Jan;2(1):31–41. doi: 10.1111/j.1365-2958.1988.tb00004.x. [DOI] [PubMed] [Google Scholar]
  21. Kiino D. R., Phillips G. J., Silhavy T. J. Increased expression of the bifunctional protein PrlF suppresses overproduction lethality associated with exported beta-galactosidase hybrid proteins in Escherichia coli. J Bacteriol. 1990 Jan;172(1):185–192. doi: 10.1128/jb.172.1.185-192.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kim J., Lee Y., Kim C., Park C. Involvement of SecB, a chaperone, in the export of ribose-binding protein. J Bacteriol. 1992 Aug;174(16):5219–5227. doi: 10.1128/jb.174.16.5219-5227.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kumamoto C. A., Beckwith J. Evidence for specificity at an early step in protein export in Escherichia coli. J Bacteriol. 1985 Jul;163(1):267–274. doi: 10.1128/jb.163.1.267-274.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kumamoto C. A. Escherichia coli SecB protein associates with exported protein precursors in vivo. Proc Natl Acad Sci U S A. 1989 Jul;86(14):5320–5324. doi: 10.1073/pnas.86.14.5320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kusukawa N., Yura T., Ueguchi C., Akiyama Y., Ito K. Effects of mutations in heat-shock genes groES and groEL on protein export in Escherichia coli. EMBO J. 1989 Nov;8(11):3517–3521. doi: 10.1002/j.1460-2075.1989.tb08517.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Laloraya S., Gambill B. D., Craig E. A. A role for a eukaryotic GrpE-related protein, Mge1p, in protein translocation. Proc Natl Acad Sci U S A. 1994 Jul 5;91(14):6481–6485. doi: 10.1073/pnas.91.14.6481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Laminet A. A., Ziegelhoffer T., Georgopoulos C., Plückthun A. The Escherichia coli heat shock proteins GroEL and GroES modulate the folding of the beta-lactamase precursor. EMBO J. 1990 Jul;9(7):2315–2319. doi: 10.1002/j.1460-2075.1990.tb07403.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. 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]
  29. Lecker S., Lill R., Ziegelhoffer T., Georgopoulos C., Bassford P. J., Jr, Kumamoto C. A., Wickner W. Three pure chaperone proteins of Escherichia coli--SecB, trigger factor and GroEL--form soluble complexes with precursor proteins in vitro. EMBO J. 1989 Sep;8(9):2703–2709. doi: 10.1002/j.1460-2075.1989.tb08411.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Liberek K., Galitski T. P., Zylicz M., Georgopoulos C. The DnaK chaperone modulates the heat shock response of Escherichia coli by binding to the sigma 32 transcription factor. Proc Natl Acad Sci U S A. 1992 Apr 15;89(8):3516–3520. doi: 10.1073/pnas.89.8.3516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. 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]
  32. Phillips G. J., Silhavy T. J. Heat-shock proteins DnaK and GroEL facilitate export of LacZ hybrid proteins in E. coli. Nature. 1990 Apr 26;344(6269):882–884. doi: 10.1038/344882a0. [DOI] [PubMed] [Google Scholar]
  33. Phillips G. J., Silhavy T. J. The E. coli ffh gene is necessary for viability and efficient protein export. Nature. 1992 Oct 22;359(6397):744–746. doi: 10.1038/359744a0. [DOI] [PubMed] [Google Scholar]
  34. Sanders S. L., Schekman R. Polypeptide translocation across the endoplasmic reticulum membrane. J Biol Chem. 1992 Jul 15;267(20):13791–13794. [PubMed] [Google Scholar]
  35. Sherman MYu, Goldberg A. L. Involvement of the chaperonin dnaK in the rapid degradation of a mutant protein in Escherichia coli. EMBO J. 1992 Jan;11(1):71–77. doi: 10.1002/j.1460-2075.1992.tb05029.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Stoller G., Rücknagel K. P., Nierhaus K. H., Schmid F. X., Fischer G., Rahfeld J. U. A ribosome-associated peptidyl-prolyl cis/trans isomerase identified as the trigger factor. EMBO J. 1995 Oct 16;14(20):4939–4948. doi: 10.1002/j.1460-2075.1995.tb00177.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Straus D. B., Walter W. A., Gross C. A. Escherichia coli heat shock gene mutants are defective in proteolysis. Genes Dev. 1988 Dec;2(12B):1851–1858. doi: 10.1101/gad.2.12b.1851. [DOI] [PubMed] [Google Scholar]
  38. Straus D. B., Walter W. A., Gross C. A. The heat shock response of E. coli is regulated by changes in the concentration of sigma 32. Nature. 1987 Sep 24;329(6137):348–351. doi: 10.1038/329348a0. [DOI] [PubMed] [Google Scholar]
  39. 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]
  40. Stuart R. A., Cyr D. M., Craig E. A., Neupert W. Mitochondrial molecular chaperones: their role in protein translocation. Trends Biochem Sci. 1994 Feb;19(2):87–92. doi: 10.1016/0968-0004(94)90041-8. [DOI] [PubMed] [Google Scholar]
  41. Szabo A., Langer T., Schröder H., Flanagan J., Bukau B., Hartl F. U. The ATP hydrolysis-dependent reaction cycle of the Escherichia coli Hsp70 system DnaK, DnaJ, and GrpE. Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10345–10349. doi: 10.1073/pnas.91.22.10345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Wickner S. H. Three Escherichia coli heat shock proteins are required for P1 plasmid DNA replication: formation of an active complex between E. coli DnaJ protein and the P1 initiator protein. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2690–2694. doi: 10.1073/pnas.87.7.2690. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wickner S., Skowyra D., Hoskins J., McKenney K. DnaJ, DnaK, and GrpE heat shock proteins are required in oriP1 DNA replication solely at the RepA monomerization step. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10345–10349. doi: 10.1073/pnas.89.21.10345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Wild J., Altman E., Yura T., Gross C. A. DnaK and DnaJ heat shock proteins participate in protein export in Escherichia coli. Genes Dev. 1992 Jul;6(7):1165–1172. doi: 10.1101/gad.6.7.1165. [DOI] [PubMed] [Google Scholar]
  45. Wild J., Kamath-Loeb A., Ziegelhoffer E., Lonetto M., Kawasaki Y., Gross C. A. Partial loss of function mutations in DnaK, the Escherichia coli homologue of the 70-kDa heat shock proteins, affect highly conserved amino acids implicated in ATP binding and hydrolysis. Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):7139–7143. doi: 10.1073/pnas.89.15.7139. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wild J., Walter W. A., Gross C. A., Altman E. Accumulation of secretory protein precursors in Escherichia coli induces the heat shock response. J Bacteriol. 1993 Jul;175(13):3992–3997. doi: 10.1128/jb.175.13.3992-3997.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES