Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Apr;178(7):1962–1970. doi: 10.1128/jb.178.7.1962-1970.1996

Essential role of a sodium dodecyl sulfate-resistant protein IV multimer in assembly-export of filamentous phage.

N A Linderoth 1, P Model 1, M Russel 1
PMCID: PMC177892  PMID: 8606171

Abstract

Filamentous phage f1 encodes protein IV (pIV), a protein essential for phage morphogenesis that localizes to the outer membrane of Escherichia coli, where it is found as a multimer of 10 to 12 subunits. Introduction of internal His or Strep affinity tags at different sites in pIV interfered with its function to a variable extent. A spontaneous second-site suppressor mutation in gene IV allowed several different insertion mutants to function. The identical mutation was also isolated as a suppressor of a multimerization-defective missense mutation. A high-molecular-mass pIV species is the predominant form of pIV present in cells. This species is stable in 4% sodium dodecyl sulfate at temperatures up to 65 degrees C and is largely preserved at 100 degrees C in Laemmli protein sample buffer containing 4% sodium dodecyl sulfate. The suppressor mutation makes the high-molecular-mass form of wild-type pIV extremely resistant to dissociation, and it stabilizes the high-molecular-mass form of several mutant pIV proteins to extents that correlate with their level of function. Mixed multimers of pIV(f1) and pIV(Ike) also remain associated during heating in sodium dodecyl sulfate-containing buffers. Thus, sodium dodecyl sulfate- and heat-resistant high-molecular-mass pIV is derived from pIV multimer and reflects the physiologically relevant form of the protein essential for assembly-export.

Full Text

The Full Text of this article is available as a PDF (463.7 KB).

Selected References

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

  1. Ahmer B. M., Thomas M. G., Larsen R. A., Postle K. Characterization of the exbBD operon of Escherichia coli and the role of ExbB and ExbD in TonB function and stability. J Bacteriol. 1995 Aug;177(16):4742–4747. doi: 10.1128/jb.177.16.4742-4747.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brissette J. L., Russel M. Secretion and membrane integration of a filamentous phage-encoded morphogenetic protein. J Mol Biol. 1990 Feb 5;211(3):565–580. doi: 10.1016/0022-2836(90)90266-O. [DOI] [PubMed] [Google Scholar]
  3. Brissette J. L., Russel M., Weiner L., Model P. Phage shock protein, a stress protein of Escherichia coli. Proc Natl Acad Sci U S A. 1990 Feb;87(3):862–866. doi: 10.1073/pnas.87.3.862. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Chen L. Y., Chen D. Y., Miaw J., Hu N. T. XpsD, an outer membrane protein required for protein secretion by Xanthomonas campestris pv. campestris, forms a multimer. J Biol Chem. 1996 Feb 2;271(5):2703–2708. doi: 10.1074/jbc.271.5.2703. [DOI] [PubMed] [Google Scholar]
  6. Cowan S. W., Schirmer T., Rummel G., Steiert M., Ghosh R., Pauptit R. A., Jansonius J. N., Rosenbusch J. P. Crystal structures explain functional properties of two E. coli porins. Nature. 1992 Aug 27;358(6389):727–733. doi: 10.1038/358727a0. [DOI] [PubMed] [Google Scholar]
  7. Drake S. L., Koomey M. The product of the pilQ gene is essential for the biogenesis of type IV pili in Neisseria gonorrhoeae. Mol Microbiol. 1995 Dec;18(5):975–986. doi: 10.1111/j.1365-2958.1995.18050975.x. [DOI] [PubMed] [Google Scholar]
  8. Endemann H., Model P. Location of filamentous phage minor coat proteins in phage and in infected cells. J Mol Biol. 1995 Jul 21;250(4):496–506. doi: 10.1006/jmbi.1995.0393. [DOI] [PubMed] [Google Scholar]
  9. Fane B., Villafane R., Mitraki A., King J. Identification of global suppressors for temperature-sensitive folding mutations of the P22 tailspike protein. J Biol Chem. 1991 Jun 25;266(18):11640–11648. [PubMed] [Google Scholar]
  10. Fischer E., Günter K., Braun V. Involvement of ExbB and TonB in transport across the outer membrane of Escherichia coli: phenotypic complementation of exb mutants by overexpressed tonB and physical stabilization of TonB by ExbB. J Bacteriol. 1989 Sep;171(9):5127–5134. doi: 10.1128/jb.171.9.5127-5134.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Freudl R., Schwarz H., Stierhof Y. D., Gamon K., Hindennach I., Henning U. An outer membrane protein (OmpA) of Escherichia coli K-12 undergoes a conformational change during export. J Biol Chem. 1986 Aug 25;261(24):11355–11361. [PubMed] [Google Scholar]
  12. Genin S., Boucher C. A. A superfamily of proteins involved in different secretion pathways in gram-negative bacteria: modular structure and specificity of the N-terminal domain. Mol Gen Genet. 1994 Apr;243(1):112–118. doi: 10.1007/BF00283883. [DOI] [PubMed] [Google Scholar]
  13. Greenberger L. M., Williams S. S., Horwitz S. B. Biosynthesis of heterogeneous forms of multidrug resistance-associated glycoproteins. J Biol Chem. 1987 Oct 5;262(28):13685–13689. [PubMed] [Google Scholar]
  14. Gregory R. J., Cheng S. H., Rich D. P., Marshall J., Paul S., Hehir K., Ostedgaard L., Klinger K. W., Welsh M. J., Smith A. E. Expression and characterization of the cystic fibrosis transmembrane conductance regulator. Nature. 1990 Sep 27;347(6291):382–386. doi: 10.1038/347382a0. [DOI] [PubMed] [Google Scholar]
  15. Guy-Caffey J. K., Rapoza M. P., Jolley K. A., Webster R. E. Membrane localization and topology of a viral assembly protein. J Bacteriol. 1992 Apr;174(8):2460–2465. doi: 10.1128/jb.174.8.2460-2465.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hill D. F., Petersen G. B. Nucleotide sequence of bacteriophage f1 DNA. J Virol. 1982 Oct;44(1):32–46. doi: 10.1128/jvi.44.1.32-46.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hobbs M., Mattick J. S. Common components in the assembly of type 4 fimbriae, DNA transfer systems, filamentous phage and protein-secretion apparatus: a general system for the formation of surface-associated protein complexes. Mol Microbiol. 1993 Oct;10(2):233–243. doi: 10.1111/j.1365-2958.1993.tb01949.x. [DOI] [PubMed] [Google Scholar]
  18. Hochuli E., Döbeli H., Schacher A. New metal chelate adsorbent selective for proteins and peptides containing neighbouring histidine residues. J Chromatogr. 1987 Dec 18;411:177–184. doi: 10.1016/s0021-9673(00)93969-4. [DOI] [PubMed] [Google Scholar]
  19. Kazmierczak B. I., Mielke D. L., Russel M., Model P. pIV, a filamentous phage protein that mediates phage export across the bacterial cell envelope, forms a multimer. J Mol Biol. 1994 Apr 29;238(2):187–198. doi: 10.1006/jmbi.1994.1280. [DOI] [PubMed] [Google Scholar]
  20. Kreusch A., Schulz G. E. Refined structure of the porin from Rhodopseudomonas blastica. Comparison with the porin from Rhodobacter capsulatus. J Mol Biol. 1994 Nov 11;243(5):891–905. doi: 10.1006/jmbi.1994.1690. [DOI] [PubMed] [Google Scholar]
  21. Kuchler K., Dohlman H. G., Thorner J. The a-factor transporter (STE6 gene product) and cell polarity in the yeast Saccharomyces cerevisiae. J Cell Biol. 1993 Mar;120(5):1203–1215. doi: 10.1083/jcb.120.5.1203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  23. Lessl M., Balzer D., Lurz R., Waters V. L., Guiney D. G., Lanka E. Dissection of IncP conjugative plasmid transfer: definition of the transfer region Tra2 by mobilization of the Tra1 region in trans. J Bacteriol. 1992 Apr;174(8):2493–2500. doi: 10.1128/jb.174.8.2493-2500.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nakamura K., Mizushima S. Effects of heating in dodecyl sulfate solution on the conformation and electrophoretic mobility of isolated major outer membrane proteins from Escherichia coli K-12. J Biochem. 1976 Dec;80(6):1411–1422. doi: 10.1093/oxfordjournals.jbchem.a131414. [DOI] [PubMed] [Google Scholar]
  25. Newhall W. J., Sawyer W. D., Haak R. A. Cross-linking analysis of the outer membrane proteins of Neisseria gonorrhoeae. Infect Immun. 1980 Jun;28(3):785–791. doi: 10.1128/iai.28.3.785-791.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Newhall W. J., Wilde C. E., 3rd, Sawyer W. D., Haak R. A. High-molecular-weight antigenic protein complex in the outer membrane of Neisseria gonorrhoeae. Infect Immun. 1980 Feb;27(2):475–482. doi: 10.1128/iai.27.2.475-482.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Nikaido H. Porins and specific diffusion channels in bacterial outer membranes. J Biol Chem. 1994 Feb 11;269(6):3905–3908. [PubMed] [Google Scholar]
  28. Plano G. V., Straley S. C. Mutations in yscC, yscD, and yscG prevent high-level expression and secretion of V antigen and Yops in Yersinia pestis. J Bacteriol. 1995 Jul;177(13):3843–3854. doi: 10.1128/jb.177.13.3843-3854.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rapoza M. P., Webster R. E. The products of gene I and the overlapping in-frame gene XI are required for filamentous phage assembly. J Mol Biol. 1995 May 5;248(3):627–638. doi: 10.1006/jmbi.1995.0247. [DOI] [PubMed] [Google Scholar]
  30. Ravetch J. V., Ohsumi M., Model P., Vovis G. F., Fischhoff D., Zinder N. D. Organization of a hybrid between phage f1 and plasmid pSC101. Proc Natl Acad Sci U S A. 1979 May;76(5):2195–2198. doi: 10.1073/pnas.76.5.2195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rost B., Sander C. Improved prediction of protein secondary structure by use of sequence profiles and neural networks. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7558–7562. doi: 10.1073/pnas.90.16.7558. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Russel M. Interchangeability of related proteins and autonomy of function. The morphogenetic proteins of filamentous phage f1 and IKe cannot replace one another. J Mol Biol. 1992 Sep 20;227(2):453–462. doi: 10.1016/0022-2836(92)90900-5. [DOI] [PubMed] [Google Scholar]
  33. Russel M., Kaźmierczak B. Analysis of the structure and subcellular location of filamentous phage pIV. J Bacteriol. 1993 Jul;175(13):3998–4007. doi: 10.1128/jb.175.13.3998-4007.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Russel M., Model P. Genetic analysis of the filamentous bacteriophage packaging signal and of the proteins that interact with it. J Virol. 1989 Aug;63(8):3284–3295. doi: 10.1128/jvi.63.8.3284-3295.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Russel M., Model P. Thioredoxin is required for filamentous phage assembly. Proc Natl Acad Sci U S A. 1985 Jan;82(1):29–33. doi: 10.1073/pnas.82.1.29. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Russel M. Moving through the membrane with filamentous phages. Trends Microbiol. 1995 Jun;3(6):223–228. doi: 10.1016/s0966-842x(00)88929-5. [DOI] [PubMed] [Google Scholar]
  37. Russel M. Mutants at conserved positions in gene IV, a gene required for assembly and secretion of filamentous phages. Mol Microbiol. 1994 Oct;14(2):357–369. doi: 10.1111/j.1365-2958.1994.tb01296.x. [DOI] [PubMed] [Google Scholar]
  38. Russel M. Phage assembly: a paradigm for bacterial virulence factor export? Science. 1994 Jul 29;265(5172):612–614. doi: 10.1126/science.8036510. [DOI] [PubMed] [Google Scholar]
  39. Russel M. Protein-protein interactions during filamentous phage assembly. J Mol Biol. 1993 Jun 5;231(3):689–697. doi: 10.1006/jmbi.1993.1320. [DOI] [PubMed] [Google Scholar]
  40. Sahin-Tóth M., Kaback H. R. Cysteine scanning mutagenesis of putative transmembrane helices IX and X in the lactose permease of Escherichia coli. Protein Sci. 1993 Jun;2(6):1024–1033. doi: 10.1002/pro.5560020615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Schirmer T., Keller T. A., Wang Y. F., Rosenbusch J. P. Structural basis for sugar translocation through maltoporin channels at 3.1 A resolution. Science. 1995 Jan 27;267(5197):512–514. doi: 10.1126/science.7824948. [DOI] [PubMed] [Google Scholar]
  42. Schmidt T. G., Skerra A. The random peptide library-assisted engineering of a C-terminal affinity peptide, useful for the detection and purification of a functional Ig Fv fragment. Protein Eng. 1993 Jan;6(1):109–122. doi: 10.1093/protein/6.1.109. [DOI] [PubMed] [Google Scholar]
  43. Sen K., Nikaido H. In vitro trimerization of OmpF porin secreted by spheroplasts of Escherichia coli. Proc Natl Acad Sci U S A. 1990 Jan;87(2):743–747. doi: 10.1073/pnas.87.2.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Shortle D., Lin B. Genetic analysis of staphylococcal nuclease: identification of three intragenic "global" suppressors of nuclease-minus mutations. Genetics. 1985 Aug;110(4):539–555. doi: 10.1093/genetics/110.4.539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Taura T., Baba T., Akiyama Y., Ito K. Determinants of the quantity of the stable SecY complex in the Escherichia coli cell. J Bacteriol. 1993 Dec;175(24):7771–7775. doi: 10.1128/jb.175.24.7771-7775.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. VOGEL H. J., BONNER D. M. Acetylornithinase of Escherichia coli: partial purification and some properties. J Biol Chem. 1956 Jan;218(1):97–106. [PubMed] [Google Scholar]
  47. Weiner L., Brissette J. L., Model P. Stress-induced expression of the Escherichia coli phage shock protein operon is dependent on sigma 54 and modulated by positive and negative feedback mechanisms. Genes Dev. 1991 Oct;5(10):1912–1923. doi: 10.1101/gad.5.10.1912. [DOI] [PubMed] [Google Scholar]
  48. Zinder N. D., Boeke J. D. The filamentous phage (Ff) as vectors for recombinant DNA--a review. Gene. 1982 Jul-Aug;19(1):1–10. doi: 10.1016/0378-1119(82)90183-4. [DOI] [PubMed] [Google Scholar]

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

RESOURCES