1887

Abstract

encodes a major phase-variable outer-membrane protein, antigen 43 (Ag43), involved in autoaggregation of cells. The gene is present in single copy on the chromosome of K-12. In contrast, Southern hybridization and gene inactivation studies demonstrate that control producer strain ML308-225 possesses duplicate copies of ( and ). Construction and analyses of single and double knockout mutants clearly show that both alleles are capable of expressing antigen in a phase-variable manner, with observed differences in the ON↔OFF switch frequencies appearing to favour expression of Ag43B under conditions of normal laboratory growth. Comparative analysis of and gene sequences revealed 98% identity at the nucleotide and predicted protein levels, with differences in the protein sequence of the surface-expressed α subunit altering the surface probability of one of the predicted epitopes. Analysis of a panel of enteropathogenic strains by Southern hybridization using -specific gene probes provided strong evidence for the presence of varying numbers of alleles within clinical isolates. Taken together, the results indicate the presence of a family of distinct Ag43 proteins encoded by multiple chromosomal alleles.

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2001-01-01
2019-10-21
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References

  1. Barbour, A. G. ( 1993; ). Linear DNA of Borrelia species and antigenic variation. Trends Microbiol 1, 236-239.[CrossRef]
    [Google Scholar]
  2. Blattner, F. R., Plunkett, G., 3rd, Bloch, C. A. & 14 other authors ( 1997; ). The complete genome sequence of Escherichia coli K-12. Science 277, 1453–1462.[CrossRef]
    [Google Scholar]
  3. Blattner, F. R., Burland, V., Durfee, T. & 10 other authors (2000). http://www.genome.wisc.edu/.
  4. Brimer, C. D. & Montie, T. C. ( 1998; ). Cloning and comparison of fliC genes and identification of glycosylation in the flagellin of Pseudomonas aeruginosa a-type strains. J Bacteriol 180, 3209-3217.
    [Google Scholar]
  5. Caffrey, P. & Owen, P. ( 1989; ). Purification and N-terminal sequence of the α subunit of Antigen 43, a unique protein complex associated with the outer membrane of Escherichia coli. J Bacteriol 171, 3634-3640.
    [Google Scholar]
  6. Caffrey, P., McVeigh, T. & Owen, P. ( 1988; ). Western immunoblotting. In Immunochemical and Molecular Genetic Analysis of Bacterial Pathogens , pp. 255-266. Edited by P. Owen & T. J. Foster. Amsterdam:Elsevier.
  7. Danese, P. N., Pratt, L. A., Dove, S. L. & Kolter, R. ( 2000; ). The outer membrane protein, Antigen 43, mediates cell-to-cell interactions within Escherichia coli biofilms. Mol Microbiol 37, 424-432.[CrossRef]
    [Google Scholar]
  8. Dehio, C., Gray-Owen, S. D. & Meyer, T. F. ( 1998; ). The role of neisserial Opa proteins in interactions with host cells. Trends Microbiol 6, 489-495.[CrossRef]
    [Google Scholar]
  9. Diderichsen, B. ( 1980; ). flu, a metastable gene controlling surface properties of Escherichia coli. J Bacteriol 141, 858-867.
    [Google Scholar]
  10. Eisenstein, B. I. ( 1981; ). Phase variation of type 1 fimbriae in Escherichia coli is under transcriptional control. Science 214, 337-339.[CrossRef]
    [Google Scholar]
  11. Feliciello, I. & Chinali, G. ( 1993; ). A modified alkaline lysis method for the preparation of highly purified plasmid DNA from Escherichia coli. Anal Biochem 212, 394-401.[CrossRef]
    [Google Scholar]
  12. Force, A., Lynch, M., Pickett, F. B., Amores, A., Yan, Y. L. & Postlewait, J. ( 1999; ). Preservation of duplicate genes by complementary, degenerative mutations. Genetics 151, 1531-1545.
    [Google Scholar]
  13. Forest, K. T., Dunham, S. A., Koomey, M. & Tainer, J. A. ( 1999; ). Crystallographic structure reveals phosphorylated pilin from Neisseria: phosphoserine sites modify type IV pilus surface chemistry and fibre morphology. Mol Microbiol 31, 743-752.[CrossRef]
    [Google Scholar]
  14. Haagmans, W. & van der Woude, M. ( 2000; ). Phase variation of Ag43 in Escherichia coli: Dam-dependent methylation abrogates OxyR binding and OxyR-mediated repression of transcription. Mol Microbiol 35, 877-887.[CrossRef]
    [Google Scholar]
  15. Hacker, J., Blum-Oehler, G., Muhldorfer, I. & Tschape, H. ( 1997; ). Pathogenicity islands of virulent bacteria: structure, function and impact on microbial evolution. Mol Microbiol 23, 1089-1097.[CrossRef]
    [Google Scholar]
  16. Hasman, H., Chakraborty, T. & Klemm, P. ( 1999; ). Antigen-43-mediated autoaggregation of Escherichia coli is blocked by fimbriation. J Bacteriol 181, 4834-4841.
    [Google Scholar]
  17. Hasman, H., Schembri, M. A. & Klemm, P. ( 2000; ). Antigen 43 and type 1 fimbriae determine colony morphology of Escherichia coli K-12. J Bacteriol 182, 1089-1095.[CrossRef]
    [Google Scholar]
  18. Henderson, I. R. & Owen, P. ( 1999; ). The major phase-variable outer membrane protein of Escherichia coli structurally resembles the immunoglobulin A1 protease class of exported protein and is regulated by a novel mechanism involving Dam and OxyR. J Bacteriol 181, 2132-2141.
    [Google Scholar]
  19. Henderson, I. R., Meehan, M. & Owen, P. ( 1997a; ). Antigen 43, a phase-variable bipartite outer membrane protein, determines colony morphology and autoaggregation in Escherichia coli K-12. FEMS Microbiol Lett 149, 115-120.[CrossRef]
    [Google Scholar]
  20. Henderson, I. R., Meehan, M. & Owen, P. ( 1997b; ). A novel regulatory mechanism for a novel phase-variable outer membrane protein of Escherichia coli. Adv Exp Med Biol 412, 349-355.
    [Google Scholar]
  21. Henderson, I. R., Navarro-Garcia, F. & Nataro, J. P. ( 1998; ). The great escape: structure and function of the autotransporter proteins. Trends Microbiol 6, 370-378.[CrossRef]
    [Google Scholar]
  22. Jameson, B. A. & Wolf, H. ( 1988; ). The antigenic index: a novel algorithm for predicting antigenic determinants. Comput Appl Biosci 4, 181-186.
    [Google Scholar]
  23. Kaback, H. R. ( 1971; ). Bacterial membranes. Methods Enzymol 22, 99-120.
    [Google Scholar]
  24. Kaback, H. R. ( 1976; ). Molecular biology and energetics of membrane transport. J Cell Physiol 89, 575-593.[CrossRef]
    [Google Scholar]
  25. Kramer, F. M. & Coen, D. M. (1987). Enzymatic amplification of DNA by PCR: standard procedures and optimisation. In Current Protocols in Molecular Biology, pp. 15.1.1–15.1.9. Edited by F. M. Ausubel and others. New York: Wiley.
  26. Laemmli, U. K. ( 1970; ). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680-685.[CrossRef]
    [Google Scholar]
  27. Lech, K. & Brent, R. (1987). Media preparation and bacteriological tools. In Current Protocols in Molecular Biology, pp. 1.1.1–1.1.6. Edited by F. M. Ausubel and others. New York: Wiley.
  28. Lindenthal, C. & Elsinghorst, E. A. ( 1999; ). Identification of a glycoprotein produced by enterotoxigenic Escherichia coli. Infect Immun 67, 4084-4091.
    [Google Scholar]
  29. Maurer, J., Jose, J. & Meyer, T. F. ( 1999; ). Characterization of the essential transport function of the AIDA-I autotransporter and evidence supporting structural predictions. J Bacteriol 181, 7014-7020.
    [Google Scholar]
  30. Owen, P. ( 1983; ). Antigens of the Escherichia coli cell envelope. In Electroimmunochemical Analysis of Membrane Proteins , pp. 347-373. Edited by O. J. Bjerrum. Amsterdam:Elsevier.
  31. Owen, P. ( 1986; ). An improved procedure for polypeptide analysis of radiolabeled antigens resolved by crossed immunoelectrophoresis and its application to the study of inner and outer membranes of Escherichia coli. Electrophoresis 6, 19-28.
    [Google Scholar]
  32. Owen, P. & Kaback, H. R. ( 1978; ). Molecular structure of membrane vesicles from Escherichia coli. Proc Natl Acad Sci USA 75, 3148-3152.[CrossRef]
    [Google Scholar]
  33. Owen, P., Caffery, P. & Josefsson, L.-G. ( 1987; ). Identification and partial characterization of a novel bipartite protein antigen associated with the outer membrane of Escherichia coli. J Bacteriol 169, 3770-3777.
    [Google Scholar]
  34. Owen, P., Meehan, M., de Loughry-Doherty, H. & Henderson, I. R. ( 1996; ). Phase variable outer membrane proteins in Escherichia coli. FEMS Immunol Med Microbiol 16, 63-76.[CrossRef]
    [Google Scholar]
  35. Phizicky, E. M. & Fields, S. ( 1995; ). Protein–protein interactions: methods for detection and analysis. Microbiol Rev 59, 94-123.
    [Google Scholar]
  36. Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  37. Schmitt, C. K., McKee, M. L. & O’Brien, A. D. ( 1991; ). Two copies of Shiga-like toxin II-related genes common in enterohemorrhagic Escherichia coli strains are responsible for the antigenic heterogeneity of the O157:H strain E32511. Infect Immun 59, 1065-1073.
    [Google Scholar]
  38. Sleytr, U. B. & Beveridge, T. J. ( 1999; ). Bacterial S-layers. Trends Microbiol 7, 253-260.[CrossRef]
    [Google Scholar]
  39. Stimson, E., Virji, M., Makepeace, K. & 9 other authors ( 1995; ). Meningococcal pilin: a glycoprotein substituted with digalactosyl 2,4-diacetamido-2,4,6-trideoxyhexose. Mol Microbiol 17, 1201–1214.[CrossRef]
    [Google Scholar]
  40. Wilson, K. (1987). Preparation of genomic DNA from bacteria. In Current Protocols in Molecular Biology, pp. 2.4.1–2.4.5. Edited by F. M. Ausubel and others. New York: Wiley.
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