Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1986 Jan;51(1):233–239. doi: 10.1128/iai.51.1.233-239.1986

A defined fragment of bacterial protein I (OmpF) is a polyclonal B-cell activator.

M Vordermeier, K Stäb, W G Bessler
PMCID: PMC261092  PMID: 3484458

Abstract

Protein I from the outer membrane of Escherichia coli and other members of the family Enterobacteriaceae is a potent mitogen and polyclonal B-lymphocyte activator. To determine the part of the polypeptide responsible for biological activity, we cleaved the molecule into defined polypeptide fragments of approximate molecular weights 24,000, 15,000, 9,000, 7,000, and 3,000 by using the cyanogen bromide method. The fragments were purified by gel permeation chromatography and by preparative polyacrylamide gel electrophoresis. They were investigated for mitogenicity and for the induction of immunoglobulin synthesis in lymphocyte cultures from several inbred mouse strains. The fragment of molecular weight 24,000 turned out to be a potent polyclonal B-lymphocyte activator comparable to native protein I. The low-molecular-weight fragments exhibited only marginal effects. Neither purified T lymphocytes nor thymocytes were activated. Our results show that a defined fragment of protein I is responsible for its lymphocyte-stimulating activity.

Full text

PDF
233

Images in this article

234Image
on p.234
235Image
on p.235

Selected References

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

  1. Andersson J., Melchers F., Galanos C., Lüderitz O. The mitogenic effect of lipopolysaccharide on bone marrow-derived mouse lymphocytes. Lipid A as the mitogenic part of the molecule. J Exp Med. 1973 Apr 1;137(4):943–953. doi: 10.1084/jem.137.4.943. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Andersson J., Sjöberg O., Möller G. Induction of immunoglobulin and antibody synthesis in vitro by lipopolysaccharides. Eur J Immunol. 1972 Aug;2(4):349–353. doi: 10.1002/eji.1830020410. [DOI] [PubMed] [Google Scholar]
  3. Bessler W. G., Cox M., Wiesmüller K. H., Jung G. The mitogenic principle of Escherichia coli lipoprotein: B-lymphocyte mitogenicity of the synthetic analogue palmitoyl-tetrapeptide (Pam-Ser-Ser-Asn-Ala). Biochem Biophys Res Commun. 1984 May 31;121(1):55–61. doi: 10.1016/0006-291x(84)90687-9. [DOI] [PubMed] [Google Scholar]
  4. Bessler W. G., Henning U. Protein I and protein II from the outer membrane of Escherichia coli are mouse B-lymphocyte mitogens. Z Immunitatsforsch Immunobiol. 1979 Jun;155(5):387–398. [PubMed] [Google Scholar]
  5. Bessler W. G., Johnson R. B., Wiesmüller K., Jung G. B-lymphocyte mitogenicity in vitro of a synthetic lipopeptide fragment derived from bacterial lipoprotein. Hoppe Seylers Z Physiol Chem. 1982 Jul;363(7):767–770. [PubMed] [Google Scholar]
  6. Bessler W., Resch K., Hancock E., Hantke K. Induction of lymphocyte proliferation and membrane changes by lipopeptide derivatives of the lipoprotein from the outer membrane of Escherichia coli. Z Immunitatsforsch Immunobiol. 1977 Apr;153(1):11–22. [PubMed] [Google Scholar]
  7. Chen R., Hindennach I., Henning U. Major proteins of the Escherichia coli outer cell envelope membrane. Sequence of the cyanogen bromide fragments of protein I from Escherichia coli B/r. Hoppe Seylers Z Physiol Chem. 1978 Dec;359(12):1807–1810. [PubMed] [Google Scholar]
  8. Chen R., Krämer C., Schmidmayr W., Chen-Schmeisser U., Henning U. Primary structure of major outer-membrane protein I (ompF protein, porin) of Escherichia coli B/r. Biochem J. 1982 Apr 1;203(1):33–43. doi: 10.1042/bj2030033. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chen R., Krämer C., Schmidmayr W., Henning U. Primary structure of major outer membrane protein I of Escherichia coli B/r. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5014–5017. doi: 10.1073/pnas.76.10.5014. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Coutinho A., Forni L., Watanabe T. Genetic and functional characterization of an antiserum to the lipid A-specific triggering receptor on murine B lymphocytes. Eur J Immunol. 1978 Jan;8(1):63–67. doi: 10.1002/eji.1830080113. [DOI] [PubMed] [Google Scholar]
  11. Dixon H. B., Perham R. N. Reversible blocking of amino groups with citraconic anhydride. Biochem J. 1968 Sep;109(2):312–314. doi: 10.1042/bj1090312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gamon K., Chen R., Henning U. Major proteins of the outer cell envelope membrane of Escherichia coli K12: multiple species of protein I differ in primary structure. Mol Gen Genet. 1978 Oct 30;166(2):187–192. doi: 10.1007/BF00285921. [DOI] [PubMed] [Google Scholar]
  13. Gronowicz E., Coutinho A., Melchers F. A plaque assay for all cells secreting Ig of a given type or class. Eur J Immunol. 1976 Aug;6(8):588–590. doi: 10.1002/eji.1830060812. [DOI] [PubMed] [Google Scholar]
  14. Hashimoto F., Horigome T., Kanbayashi M., Yoshida K., Sugano H. An improved method for separation of low-molecular-weight polypeptides by electrophoresis in sodium dodecyl sulfate-polyacrylamide gel. Anal Biochem. 1983 Feb 15;129(1):192–199. doi: 10.1016/0003-2697(83)90068-4. [DOI] [PubMed] [Google Scholar]
  15. Hindennach I., Henning U. The major proteins of the Excherichia coli outer cell envelope membrane. Preparative isolation of all major membrane proteins. Eur J Biochem. 1975 Nov 1;59(1):207–213. doi: 10.1111/j.1432-1033.1975.tb02443.x. [DOI] [PubMed] [Google Scholar]
  16. Johnson R. B., Köhl S., Wiesmüller K., Jung G., Bessler W. G. Synthetic analogues of the N-terminal lipid part of bacterial lipoprotein are B-lymphocyte mitogens in vitro and in vivo. Immunobiology. 1983 Jul;165(1):27–35. doi: 10.1016/S0171-2985(83)80044-8. [DOI] [PubMed] [Google Scholar]
  17. Kotani S., Takada H., Tsujimoto M., Ogawa T., Harada K., Mori Y., Kawasaki A., Tanaka A., Nagao S., Tanaka S. Immunobiologically active lipid A analogs synthesized according to a revised structural model of natural lipid A. Infect Immun. 1984 Jul;45(1):293–296. doi: 10.1128/iai.45.1.293-296.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. Melchers F., Braun V., Galanos C. The lipoprotein of the outer membrane of Escherichia coli: a B-lymphocyte mitogen. J Exp Med. 1975 Aug 1;142(2):473–482. doi: 10.1084/jem.142.2.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mohri S., Watanabe T., Nariuchi H. Studies of the immunological activities of the outer membrane protein from Escherichia coli. Immunology. 1982 Jun;46(2):271–280. [PMC free article] [PubMed] [Google Scholar]
  21. Morrissey J. H. Silver stain for proteins in polyacrylamide gels: a modified procedure with enhanced uniform sensitivity. Anal Biochem. 1981 Nov 1;117(2):307–310. doi: 10.1016/0003-2697(81)90783-1. [DOI] [PubMed] [Google Scholar]
  22. Peavy D. L., Baughn R. E., Musher D. M. Mitogenic activity of bacterial lipopolysaccharides in vivo: morphological and functinal characterization of responding cells. Infect Immun. 1978 Jan;19(1):71–78. doi: 10.1128/iai.19.1.71-78.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rittenberg M. B., Pratt K. L. Antitrinitrophenyl (TNP) plaque assay. Primary response of Balb/c mice to soluble and particulate immunogen. Proc Soc Exp Biol Med. 1969 Nov;132(2):575–581. doi: 10.3181/00379727-132-34264. [DOI] [PubMed] [Google Scholar]
  24. Schroeder W. A., Shelton J. B., Shelton J. R. An examination of conditions for the cleavage of polypeptide chains with cyanogen bromide: application to catalase. Arch Biochem Biophys. 1969 Mar;130(1):551–556. doi: 10.1016/0003-9861(69)90069-1. [DOI] [PubMed] [Google Scholar]
  25. Sultzer B. M., Nilsson B. S. PPD tuberculin--a B-cell mitogen. Nat New Biol. 1972 Dec 13;240(102):198–200. doi: 10.1038/newbio240198a0. [DOI] [PubMed] [Google Scholar]
  26. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  27. Wiesmüller K. H., Bessler W., Jung G. Synthesis of the mitogenic S-[2,3-bis(palmitoyloxy)propyl]-N-palmitoylpentapeptide from Escherichia coli lipoprotein. Hoppe Seylers Z Physiol Chem. 1983 May;364(5):593–606. doi: 10.1515/bchm2.1983.364.1.593. [DOI] [PubMed] [Google Scholar]
  28. Yokoyama K., Mashimo J., Kasai N., Terao T., Osawa T. Binding of bacterial lipopolysaccharide to histocompatibility-2-complex proteins of mouse lymphocytes. Hoppe Seylers Z Physiol Chem. 1979 Apr;360(4):587–595. doi: 10.1515/bchm2.1979.360.1.587. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES