Molecular analysis of VcfQ protein involved in type IV pilus biogenesis Free

Abstract

The nucleotide sequence of an ORF () within the type IV pilus gene cluster of O34 strain NAGV14 was determined, thereby completing the sequence analysis of the structural operon. The gene showed homology to the gene of the mannose-sensitive haemagglutinin pilus gene cluster. The was 651 bp larger than , and the G+C content of the extra 651 bp portion (35.6 mol%) was lower than that of the overall gene (42.5 mol%). Except for the first 270 aa residues, the deduced amino acid sequence of VcfQ showed high homology to the MshQ protein. There was immunological cross-reaction between VcfQ and MshQ by Western blotting. Cell fractionation studies showed that VcfQ is located in both the inner and the outer membranes. Mutational analysis showed that -deficient mutant expressed detectable levels of major pilin (VcfA), but failed to assemble them into pili, indicating that VcfQ is essential for pilus assembly. Colony-blotting analyses showed that the N-terminal region of is variable in strains.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.04967-0
2003-04-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/jmm/52/4/283.html?itemId=/content/journal/jmm/10.1099/jmm.0.04967-0&mimeType=html&fmt=ahah

References

  1. Alm R. A, Bina J, Andrews B. M, Doig P, Hancock R. E. W, Trust T. J. 2000; Comparative genomics of Helicobacter pylori : analysis of the outer membrane protein families. Infect Immun 68:4155–4168 [CrossRef]
    [Google Scholar]
  2. Bagdasarian M, Bagdasarian M. M. 1994; Gene cloning and expression. In Methods for General and Molecular Bacteriology pp 409–412 Edited by Gerhardt P., Murray R. G. E., Wood W. A., Krieg N. R. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  3. Bhattacharya M. K, Dutta D, Bhattacharya S. K. 7 other authors 1998; Association of a disease approximating cholera caused by Vibrio cholerae of serogroups other than O1 and O139. Epidemiol Infect 120:1–5 [CrossRef]
    [Google Scholar]
  4. Birnboim H. C, Doly J. 1979; A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523 [CrossRef]
    [Google Scholar]
  5. Castric P. 1995; pilO , a gene required for glycosylation of Pseudomonas aeruginosa 1244 pilin. Microbiology 141:1247–1254 [CrossRef]
    [Google Scholar]
  6. Chiavelli D. A, Marsh J. W, Taylor R. K. 2001; The mannose-sensitive hemagglutinin of Vibrio cholerae promotes adherence to zooplankton. Appl Environ Microbiol 67:3220–3225 [CrossRef]
    [Google Scholar]
  7. Dalsgaard A, Forslund A, Bodhidatta L, Serichantalergs O, Pitarangsi C, Pang L, Shimada T, Echeverria P. 1999; A high proportion of Vibrio cholerae strains isolated from children with diarrhea in Bangkok, Thailand are multiple antibiotic resistant and belong to heterogeneous non-O1, non-O139 O-serotypes. Epidemiol Infect 122:217–226 [CrossRef]
    [Google Scholar]
  8. Faruque S. M, Albert M. J, Mekalanos J. J. 1998; Epidemiology, genetics, and ecology of toxigenic Vibrio cholerae . Microbiol Mol Biol Rev 62:1301–1314
    [Google Scholar]
  9. Iwanaga M, Nakasone N, Ehara M. 1989; A comparative study on adhesive and non-adhesive strains. Microbiol Immunol 33:1–9 [CrossRef]
    [Google Scholar]
  10. Karaolis D. K. R, Somara S, Maneval D. R Jr, Johnson J. A, Kaper J. B. 1999; A bacteriophage encoding a pathogenicity island, a type IV pilus and a phage receptor in cholera bacteria. Nature 399:375–379 [CrossRef]
    [Google Scholar]
  11. Kuroki H, Toma C, Nakasone N, Yamashiro T, Iwanaga M. 2001; Gene analysis of Vibrio cholerae NAGV14 pilus and its distribution. Microbiol Immunol 45:417–424 [CrossRef]
    [Google Scholar]
  12. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [CrossRef]
    [Google Scholar]
  13. Lawrence J. G, Ochman H. 1998; Molecular archaeology of the Escherichia coli genome. Proc Natl Acad Sci U S A 95:9413–9417 [CrossRef]
    [Google Scholar]
  14. Marsh J. W, Taylor R. K. 1999; Genetic and transcriptional analysis of the Vibrio cholerae mannose-sensitive hemagglutinin type 4 pilus gene locus. J Bacteriol 181:1110–1117
    [Google Scholar]
  15. Nakai K, Kanehisa M. 1991; Expert system for predicting protein localization sites in gram-negative bacteria. Proteins 11:95–110 [CrossRef]
    [Google Scholar]
  16. Nakasone N, Iwanaga M. 1990; Pili of Vibrio cholerae non-O1. Infect Immun 58:1640–1646
    [Google Scholar]
  17. Nakasone N, Iwanaga M. 1998; Characterization of outer membrane protein OmpU of Vibrio cholerae O1. Infect Immun 66:4726–4728
    [Google Scholar]
  18. Power P. M, Roddam L. F, Dieckelmann M, Srikhanta Y. N, Tan Y. C, Berrington A. W, Jennings M. P. 2002; Genetic characterization of pilin glycosylation in Neisseria meningitidis . Microbiology 146:967–979
    [Google Scholar]
  19. Ramer S. W, Schoolink G. K, Wu C. Y, Hwang J, Schmidt S. A, Bieber D. 2002; The type IV pilus assembly complex: biogenic interactions among the bundle-forming pilus proteins of enteropathogenic Escherichia coli . J Bacteriol 184:3457–3465 [CrossRef]
    [Google Scholar]
  20. Sharma C, Thungapathra M, Ghosh A. 11 other authors 1998; Molecular analysis of non-O1, non-O139 Vibrio cholerae associated with an unusual upsurge in the incidence of cholera-like disease in Calcutta, India. J Clin Microbiol 36:756–763
    [Google Scholar]
  21. Simon R, Priefer U, Pühler A. 1983; A broad host range mobilization system for in vivo genetic engineering: transposon mutagenesis in gram negative bacteria. Bio-technology 11:784–791
    [Google Scholar]
  22. Strom M. S, Lory S. 1993; Structure-function and biogenesis of the type IV pili. Annu Rev Microbiol 47:565–596 [CrossRef]
    [Google Scholar]
  23. Taylor R. K, Miller V. L, Furlong D. B, Mekalanos J. J. 1987; Use of phoA gene fusions to identify a pilus colonization factor coordinately regulated with cholerae toxin. Proc Natl Acad Sci U S A 84:2833–2837 [CrossRef]
    [Google Scholar]
  24. Toma C, Kuroki H, Nakasone N, Ehara M, Iwanaga M. 2002; Minor pilin subunits are conserved in Vibrio cholerae type IV pili. FEMS Immunol Med Microbiol 33:35–40 [CrossRef]
    [Google Scholar]
  25. Towbin H, Staehelin T, Gordon J. 1979; Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76:4350–4354 [CrossRef]
    [Google Scholar]
  26. Watnick P. I, Kolter R. 1999; Steps in the development of a Vibrio cholerae El Tor biofilm. Mol Microbiol 34:586–595 [CrossRef]
    [Google Scholar]
  27. Whayeb L. A, Yamamoto K, Castillo M. E. Y, Tojo H, Honda T. 1996; Lysophospholipase L2 of Vibrio cholerae O1 affects cholera toxin production. FEMS Immunol Med Microbiol 15:9–15 [CrossRef]
    [Google Scholar]
  28. Yamai S, Okitsu T, Shimada T, Katsube Y. 1997; Distribution of serogroups Vibrio cholerae non-O1 non-O139 with specific reference to their ability to produce cholera toxin and additional of novel serogroups. Jpn J Assoc Infect Dis 71:1037–1045
    [Google Scholar]
  29. Yamashiro T, Iwanaga M. 1996; Purification and characterization of a pilus of a Vibrio cholerae strain: a possible colonization factor. Infect Immun 64:5233–5238
    [Google Scholar]
  30. Yamashiro T, Nakasone N, Iwanaga M. 1993; Purification and characterization of pili of a Vibrio cholerae non-O1 strain. Infect Immun 61:5398–5400
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.04967-0
Loading
/content/journal/jmm/10.1099/jmm.0.04967-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Supplementary material 2

PDF

Most cited Most Cited RSS feed