Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1990 Sep;172(9):5307–5311. doi: 10.1128/jb.172.9.5307-5311.1990

Associations of Escherichia coli K-12 OmpF trimers with rough and smooth lipopolysaccharides.

D L Diedrich 1, M A Stein 1, C A Schnaitman 1
PMCID: PMC213194  PMID: 2168378

Abstract

The associations of both rough and smooth lipopolysaccharides (LPS) with the OmpF porin of Escherichia coli K-12 were examined in galE strains deleted for ompC. Transformation with pSS37 and growth with galactose conferred the ability to assemble a Shigella dysenteriae O antigen onto the core oligosaccharide of E. coli K-12 LPS. The association of LPS with OmpF trimers was assessed by staining, autoradiography of LPS specifically labeled with [1-14C]galactose, and Western immunoblotting with a monoclonal antibody specific for OmpF trimers. These techniques revealed that the migration distances and multiple banding patterns of OmpF porin trimers in sodium dodecyl sulfate-polyacrylamide gels were dictated by the chemotype of associated LPS. Expression of smooth LPS caused almost all of the trimeric OmpF to run in gels with a slower mobility than trimers from rough strains. The LPS associated with trimers from a smooth strain differed from the bulk-phase LPS by consisting almost exclusively of molecules with O antigen.

Full text

PDF
5307

Images in this article

5308Image
on p.5308
5308Image
on p.5308
5309Image
on p.5309
5309Image
on p.5309
5310Image
on p.5310
5310Image
on p.5310
5310Image
on p.5310

Selected References

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

  1. Benz R., Bauer K. Permeation of hydrophilic molecules through the outer membrane of gram-negative bacteria. Review on bacterial porins. Eur J Biochem. 1988 Sep 1;176(1):1–19. doi: 10.1111/j.1432-1033.1988.tb14245.x. [DOI] [PubMed] [Google Scholar]
  2. Borneleit P., Blechschmidt B., Kleber H. P. Lipopolysaccharide-protein interactions: determination of dissociation constants by affinity electrophoresis. Electrophoresis. 1989 Dec;10(12):848–852. doi: 10.1002/elps.1150101209. [DOI] [PubMed] [Google Scholar]
  3. Dunn S. D. Effects of the modification of transfer buffer composition and the renaturation of proteins in gels on the recognition of proteins on Western blots by monoclonal antibodies. Anal Biochem. 1986 Aug 15;157(1):144–153. doi: 10.1016/0003-2697(86)90207-1. [DOI] [PubMed] [Google Scholar]
  4. Gehring K. B., Nikaido H. Existence and purification of porin heterotrimers of Escherichia coli K12 OmpC, OmpF, and PhoE proteins. J Biol Chem. 1989 Feb 15;264(5):2810–2815. [PubMed] [Google Scholar]
  5. Goldman R. C., Leive L. Heterogeneity of antigenic-side-chain length in lipopolysaccharide from Escherichia coli 0111 and Salmonella typhimurium LT2. Eur J Biochem. 1980;107(1):145–153. doi: 10.1111/j.1432-1033.1980.tb04635.x. [DOI] [PubMed] [Google Scholar]
  6. Hodgson A. L., Bird P., Nisbet I. T. Cloning, nucleotide sequence, and expression in Escherichia coli of the phospholipase D gene from Corynebacterium pseudotuberculosis. J Bacteriol. 1990 Mar;172(3):1256–1261. doi: 10.1128/jb.172.3.1256-1261.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Holzenburg A., Engel A., Kessler R., Manz H. J., Lustig A., Aebi U. Rapid isolation of OmpF porin-LPS complexes suitable for structure-function studies. Biochemistry. 1989 May 16;28(10):4187–4193. doi: 10.1021/bi00436a010. [DOI] [PubMed] [Google Scholar]
  8. Jiao B. H., Freudenberg M., Galanos C. Characterization of the lipid A component of genuine smooth-form lipopolysaccharide. Eur J Biochem. 1989 Apr 1;180(3):515–518. doi: 10.1111/j.1432-1033.1989.tb14676.x. [DOI] [PubMed] [Google Scholar]
  9. Johnston K. H., Zabriskie J. B. Purification and partial characterization of the nephritis strain-associated protein from Streptococcus pyogenes, group A. J Exp Med. 1986 Mar 1;163(3):697–712. doi: 10.1084/jem.163.3.697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kulisek E. S., Holm S. E., Johnston K. H. A chromogenic assay for the detection of plasmin generated by plasminogen activator immobilized on nitrocellulose using a para-nitroanilide synthetic peptide substrate. Anal Biochem. 1989 Feb 15;177(1):78–84. doi: 10.1016/0003-2697(89)90017-1. [DOI] [PubMed] [Google Scholar]
  11. Lugtenberg B., Meijers J., Peters R., van der Hoek P., van Alphen L. Electrophoretic resolution of the "major outer membrane protein" of Escherichia coli K12 into four bands. FEBS Lett. 1975 Oct 15;58(1):254–258. doi: 10.1016/0014-5793(75)80272-9. [DOI] [PubMed] [Google Scholar]
  12. Nakae T. Outer membrane of Salmonella. Isolation of protein complex that produces transmembrane channels. J Biol Chem. 1976 Apr 10;251(7):2176–2178. [PubMed] [Google Scholar]
  13. Neidhardt F. C., Bloch P. L., Smith D. F. Culture medium for enterobacteria. J Bacteriol. 1974 Sep;119(3):736–747. doi: 10.1128/jb.119.3.736-747.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Palva E. T., Mäkelä P. H. Lipopolysaccharide heterogeneity in Salmonella typhimurium analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Eur J Biochem. 1980;107(1):137–143. doi: 10.1111/j.1432-1033.1980.tb04634.x. [DOI] [PubMed] [Google Scholar]
  15. Redinbaugh M. G., Turley R. B. Adaptation of the bicinchoninic acid protein assay for use with microtiter plates and sucrose gradient fractions. Anal Biochem. 1986 Mar;153(2):267–271. doi: 10.1016/0003-2697(86)90091-6. [DOI] [PubMed] [Google Scholar]
  16. Reid J., Fung H., Gehring K., Klebba P. E., Nikaido H. Targeting of porin to the outer membrane of Escherichia coli. Rate of trimer assembly and identification of a dimer intermediate. J Biol Chem. 1988 Jun 5;263(16):7753–7759. [PubMed] [Google Scholar]
  17. Rocque W. J., Coughlin R. T., McGroarty E. J. Lipopolysaccharide tightly bound to porin monomers and trimers from Escherichia coli K-12. J Bacteriol. 1987 Sep;169(9):4003–4010. doi: 10.1128/jb.169.9.4003-4010.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Schindler H., Rosenbusch J. P. Matrix protein from Escherichia coli outer membranes forms voltage-controlled channels in lipid bilayers. Proc Natl Acad Sci U S A. 1978 Aug;75(8):3751–3755. doi: 10.1073/pnas.75.8.3751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Schnaitman C. A., McDonald G. A. Regulation of outer membrane protein synthesis in Escherichia coli K-12: deletion of ompC affects expression of the OmpF protein. J Bacteriol. 1984 Aug;159(2):555–563. doi: 10.1128/jb.159.2.555-563.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Strittmatter W., Galanos C. Characterisation of protein co-extracted together with LPS in Escherichia coli, Salmonella minnesota and Yersinia enterocolitica. Microb Pathog. 1987 Jan;2(1):29–36. doi: 10.1016/0882-4010(87)90112-4. [DOI] [PubMed] [Google Scholar]
  21. Sturm S., Timmis K. N. Cloning of the rfb gene region of Shigella dysenteriae 1 and construction of an rfb-rfp gene cassette for the development of lipopolysaccharide-based live anti-dysentery vaccines. Microb Pathog. 1986 Jun;1(3):289–297. doi: 10.1016/0882-4010(86)90054-9. [DOI] [PubMed] [Google Scholar]
  22. Wollin R., Creeger E. S., Rothfield L. I., Stocker B. A., Lindberg A. A. Salmonella typhimurium mutants defective in UDP-D-galactose:lipopolysaccharide alpha 1,6-D-galactosyltransferase. Structural, immunochemical, and enzymologic studies of rfaB mutants. J Biol Chem. 1983 Mar 25;258(6):3769–3774. [PubMed] [Google Scholar]
  23. Yamada H., Mizushima S. Interaction between major outer membrane protein (O-8) and lipopolysaccharide in Escherichia coli K12. Eur J Biochem. 1980 Jan;103(1):209–218. doi: 10.1111/j.1432-1033.1980.tb04305.x. [DOI] [PubMed] [Google Scholar]

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

RESOURCES