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. 1994 Nov;176(22):6885–6891. doi: 10.1128/jb.176.22.6885-6891.1994

A plasmid-encoded prepilin peptidase gene from enteropathogenic Escherichia coli.

H Z Zhang 1, S Lory 1, M S Donnenberg 1
PMCID: PMC197057  PMID: 7961448

Abstract

Enteropathogenic Escherichia coli, a leading agent of infantile diarrhea worldwide, adheres to tissue culture cells in a pattern called "localized adherence." Localized adherence is associated with bundle-forming pili encoded by the plasmid bfpA gene, the product of which is homologous with the major structural subunit proteins of type IV fimbriae in other bacteria. Several of these proteins have been shown to be processed from a precursor by a specific prepilin peptidase. We cloned restriction fragments downstream of the bfpA gene into an E. coli-Pseudomonas aeruginosa shuttle vector and mobilized them into a P. aeruginosa prepilin peptidase (pilD) mutant. A plasmid containing a 1.3-kb PstI-BamHI fragment was able to complement the pilD mutation, as demonstrated by restoration of sensitivity to the pilus-specific bacteriophage PO4. The DNA sequence of this fragment revealed an open reading frame, designated bfpP, the predicted product of which is homologous to other prepilin peptidases, including TcpJ of Vibrio cholerae (30% identical amino acids), PulO of Klebsiella oxytoca (29%), and PilD of P. aeruginosa (28%). A bfpA::TnphoA mutant complemented with a bfpA-containing DNA fragment only partially processes the BfpA protein. When complemented with a larger fragment containing bfpP as well as bfpA, the mutant expresses the fully processed BfpA protein. P. aeruginosa PAK, but not a pilD mutant of PAK, expresses mature BfpA protein when the bfpA gene is mobilized into this strain. Thus, as in other type IV fimbria systems, enteropathogenic E. coli utilizes a specific prepilin peptidase to process the major subunit of the bundle-forming pilus. This prepilin petidase contains sequence and reciprocal functional homologies with the PilD protein of P. aeruginosa.

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Selected References

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

  1. Baldini M. M., Kaper J. B., Levine M. M., Candy D. C., Moon H. W. Plasmid-mediated adhesion in enteropathogenic Escherichia coli. J Pediatr Gastroenterol Nutr. 1983;2(3):534–538. doi: 10.1097/00005176-198302030-00023. [DOI] [PubMed] [Google Scholar]
  2. Beard M. K., Mattick J. S., Moore L. J., Mott M. R., Marrs C. F., Egerton J. R. Morphogenetic expression of Moraxella bovis fimbriae (pili) in Pseudomonas aeruginosa. J Bacteriol. 1990 May;172(5):2601–2607. doi: 10.1128/jb.172.5.2601-2607.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  4. Chikami G. K., Fierer J., Guiney D. G. Plasmid-mediated virulence in Salmonella dublin demonstrated by use of a Tn5-oriT construct. Infect Immun. 1985 Nov;50(2):420–424. doi: 10.1128/iai.50.2.420-424.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cravioto A., Tello A., Navarro A., Ruiz J., Villafán H., Uribe F., Eslava C. Association of Escherichia coli HEp-2 adherence patterns with type and duration of diarrhoea. Lancet. 1991 Feb 2;337(8736):262–264. doi: 10.1016/0140-6736(91)90868-p. [DOI] [PubMed] [Google Scholar]
  6. Donnenberg M. S., Calderwood S. B., Donohue-Rolfe A., Keusch G. T., Kaper J. B. Construction and analysis of TnphoA mutants of enteropathogenic Escherichia coli unable to invade HEp-2 cells. Infect Immun. 1990 Jun;58(6):1565–1571. doi: 10.1128/iai.58.6.1565-1571.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Donnenberg M. S., Girón J. A., Nataro J. P., Kaper J. B. A plasmid-encoded type IV fimbrial gene of enteropathogenic Escherichia coli associated with localized adherence. Mol Microbiol. 1992 Nov;6(22):3427–3437. doi: 10.1111/j.1365-2958.1992.tb02210.x. [DOI] [PubMed] [Google Scholar]
  8. Dupuy B., Pugsley A. P. Type IV prepilin peptidase gene of Neisseria gonorrhoeae MS11: presence of a related gene in other piliated and nonpiliated Neisseria strains. J Bacteriol. 1994 Mar;176(5):1323–1331. doi: 10.1128/jb.176.5.1323-1331.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dupuy B., Taha M. K., Possot O., Marchal C., Pugsley A. P. PulO, a component of the pullulanase secretion pathway of Klebsiella oxytoca, correctly and efficiently processes gonococcal type IV prepilin in Escherichia coli. Mol Microbiol. 1992 Jul;6(14):1887–1894. doi: 10.1111/j.1365-2958.1992.tb01361.x. [DOI] [PubMed] [Google Scholar]
  10. Dupuy B., Taha M. K., Pugsley A. P., Marchal C. Neisseria gonorrhoeae prepilin export studied in Escherichia coli. J Bacteriol. 1991 Dec;173(23):7589–7598. doi: 10.1128/jb.173.23.7589-7598.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Frost L. S., Carpenter M., Paranchych W. N-methylphenylalanine at the N-terminus of pilin isolated from Pseudomonas aeruginosa K. Nature. 1978 Jan 5;271(5640):87–89. doi: 10.1038/271087a0. [DOI] [PubMed] [Google Scholar]
  12. Fürste J. P., Pansegrau W., Frank R., Blöcker H., Scholz P., Bagdasarian M., Lanka E. Molecular cloning of the plasmid RP4 primase region in a multi-host-range tacP expression vector. Gene. 1986;48(1):119–131. doi: 10.1016/0378-1119(86)90358-6. [DOI] [PubMed] [Google Scholar]
  13. Girón J. A., Donnenberg M. S., Martin W. C., Jarvis K. G., Kaper J. B. Distribution of the bundle-forming pilus structural gene (bfpA) among enteropathogenic Escherichia coli. J Infect Dis. 1993 Oct;168(4):1037–1041. doi: 10.1093/infdis/168.4.1037. [DOI] [PubMed] [Google Scholar]
  14. Girón J. A., Ho A. S., Schoolnik G. K. An inducible bundle-forming pilus of enteropathogenic Escherichia coli. Science. 1991 Nov 1;254(5032):710–713. doi: 10.1126/science.1683004. [DOI] [PubMed] [Google Scholar]
  15. Gomes T. A., Blake P. A., Trabulsi L. R. Prevalence of Escherichia coli strains with localized, diffuse, and aggregative adherence to HeLa cells in infants with diarrhea and matched controls. J Clin Microbiol. 1989 Feb;27(2):266–269. doi: 10.1128/jcm.27.2.266-269.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. 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]
  17. Howard S. P., Critch J., Bedi A. Isolation and analysis of eight exe genes and their involvement in extracellular protein secretion and outer membrane assembly in Aeromonas hydrophila. J Bacteriol. 1993 Oct;175(20):6695–6703. doi: 10.1128/jb.175.20.6695-6703.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hoyne P. A., Haas R., Meyer T. F., Davies J. K., Elleman T. C. Production of Neisseria gonorrhoeae pili (fimbriae) in Pseudomonas aeruginosa. J Bacteriol. 1992 Nov;174(22):7321–7327. doi: 10.1128/jb.174.22.7321-7327.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kaufman M. R., Seyer J. M., Taylor R. K. Processing of TCP pilin by TcpJ typifies a common step intrinsic to a newly recognized pathway of extracellular protein secretion by gram-negative bacteria. Genes Dev. 1991 Oct;5(10):1834–1846. doi: 10.1101/gad.5.10.1834. [DOI] [PubMed] [Google Scholar]
  20. Koga T., Ishimoto K., Lory S. Genetic and functional characterization of the gene cluster specifying expression of Pseudomonas aeruginosa pili. Infect Immun. 1993 Apr;61(4):1371–1377. doi: 10.1128/iai.61.4.1371-1377.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lauer P., Albertson N. H., Koomey M. Conservation of genes encoding components of a type IV pilus assembly/two-step protein export pathway in Neisseria gonorrhoeae. Mol Microbiol. 1993 Apr;8(2):357–368. doi: 10.1111/j.1365-2958.1993.tb01579.x. [DOI] [PubMed] [Google Scholar]
  22. Levine M. M., Bergquist E. J., Nalin D. R., Waterman D. H., Hornick R. B., Young C. R., Sotman S. Escherichia coli strains that cause diarrhoea but do not produce heat-labile or heat-stable enterotoxins and are non-invasive. Lancet. 1978 May 27;1(8074):1119–1122. doi: 10.1016/s0140-6736(78)90299-4. [DOI] [PubMed] [Google Scholar]
  23. Levine M. M. Escherichia coli that cause diarrhea: enterotoxigenic, enteropathogenic, enteroinvasive, enterohemorrhagic, and enteroadherent. J Infect Dis. 1987 Mar;155(3):377–389. doi: 10.1093/infdis/155.3.377. [DOI] [PubMed] [Google Scholar]
  24. Levine M. M., Nataro J. P., Karch H., Baldini M. M., Kaper J. B., Black R. E., Clements M. L., O'Brien A. D. The diarrheal response of humans to some classic serotypes of enteropathogenic Escherichia coli is dependent on a plasmid encoding an enteroadhesiveness factor. J Infect Dis. 1985 Sep;152(3):550–559. doi: 10.1093/infdis/152.3.550. [DOI] [PubMed] [Google Scholar]
  25. Lipman D. J., Pearson W. R. Rapid and sensitive protein similarity searches. Science. 1985 Mar 22;227(4693):1435–1441. doi: 10.1126/science.2983426. [DOI] [PubMed] [Google Scholar]
  26. Lory S. Determinants of extracellular protein secretion in gram-negative bacteria. J Bacteriol. 1992 Jun;174(11):3423–3428. doi: 10.1128/jb.174.11.3423-3428.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mattick J. S., Bills M. M., Anderson B. J., Dalrymple B., Mott M. R., Egerton J. R. Morphogenetic expression of Bacteroides nodosus fimbriae in Pseudomonas aeruginosa. J Bacteriol. 1987 Jan;169(1):33–41. doi: 10.1128/jb.169.1.33-41.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nataro J. P., Maher K. O., Mackie P., Kaper J. B. Characterization of plasmids encoding the adherence factor of enteropathogenic Escherichia coli. Infect Immun. 1987 Oct;55(10):2370–2377. doi: 10.1128/iai.55.10.2370-2377.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Nataro J. P., Scaletsky I. C., Kaper J. B., Levine M. M., Trabulsi L. R. Plasmid-mediated factors conferring diffuse and localized adherence of enteropathogenic Escherichia coli. Infect Immun. 1985 May;48(2):378–383. doi: 10.1128/iai.48.2.378-383.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nunn D. N., Lory S. Components of the protein-excretion apparatus of Pseudomonas aeruginosa are processed by the type IV prepilin peptidase. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):47–51. doi: 10.1073/pnas.89.1.47. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nunn D. N., Lory S. Product of the Pseudomonas aeruginosa gene pilD is a prepilin leader peptidase. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3281–3285. doi: 10.1073/pnas.88.8.3281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nunn D., Bergman S., Lory S. Products of three accessory genes, pilB, pilC, and pilD, are required for biogenesis of Pseudomonas aeruginosa pili. J Bacteriol. 1990 Jun;172(6):2911–2919. doi: 10.1128/jb.172.6.2911-2919.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ogierman M. A., Zabihi S., Mourtzios L., Manning P. A. Genetic organization and sequence of the promoter-distal region of the tcp gene cluster of Vibrio cholerae. Gene. 1993 Apr 15;126(1):51–60. doi: 10.1016/0378-1119(93)90589-u. [DOI] [PubMed] [Google Scholar]
  34. Paranchych W., Frost L. S. The physiology and biochemistry of pili. Adv Microb Physiol. 1988;29:53–114. doi: 10.1016/s0065-2911(08)60346-x. [DOI] [PubMed] [Google Scholar]
  35. Pugsley A. P., Dupuy B. An enzyme with type IV prepilin peptidase activity is required to process components of the general extracellular protein secretion pathway of Klebsiella oxytoca. Mol Microbiol. 1992 Mar;6(6):751–760. doi: 10.1111/j.1365-2958.1992.tb01525.x. [DOI] [PubMed] [Google Scholar]
  36. Scaletsky I. C., Silva M. L., Trabulsi L. R. Distinctive patterns of adherence of enteropathogenic Escherichia coli to HeLa cells. Infect Immun. 1984 Aug;45(2):534–536. doi: 10.1128/iai.45.2.534-536.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sohel I., Puente J. L., Murray W. J., Vuopio-Varkila J., Schoolnik G. K. Cloning and characterization of the bundle-forming pilin gene of enteropathogenic Escherichia coli and its distribution in Salmonella serotypes. Mol Microbiol. 1993 Feb;7(4):563–575. doi: 10.1111/j.1365-2958.1993.tb01147.x. [DOI] [PubMed] [Google Scholar]
  38. Strom M. S., Lory S. Structure-function and biogenesis of the type IV pili. Annu Rev Microbiol. 1993;47:565–596. doi: 10.1146/annurev.mi.47.100193.003025. [DOI] [PubMed] [Google Scholar]
  39. Strom M. S., Nunn D. N., Lory S. A single bifunctional enzyme, PilD, catalyzes cleavage and N-methylation of proteins belonging to the type IV pilin family. Proc Natl Acad Sci U S A. 1993 Mar 15;90(6):2404–2408. doi: 10.1073/pnas.90.6.2404. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Strom M. S., Nunn D., Lory S. Multiple roles of the pilus biogenesis protein pilD: involvement of pilD in excretion of enzymes from Pseudomonas aeruginosa. J Bacteriol. 1991 Feb;173(3):1175–1180. doi: 10.1128/jb.173.3.1175-1180.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  42. Whitchurch C. B., Hobbs M., Livingston S. P., Krishnapillai V., Mattick J. S. Characterisation of a Pseudomonas aeruginosa twitching motility gene and evidence for a specialised protein export system widespread in eubacteria. Gene. 1991 May 15;101(1):33–44. doi: 10.1016/0378-1119(91)90221-v. [DOI] [PubMed] [Google Scholar]
  43. de Groot A., Filloux A., Tommassen J. Conservation of xcp genes, involved in the two-step protein secretion process, in different Pseudomonas species and other gram-negative bacteria. Mol Gen Genet. 1991 Oct;229(2):278–284. doi: 10.1007/BF00272167. [DOI] [PubMed] [Google Scholar]
  44. de Groot A., Heijnen I., de Cock H., Filloux A., Tommassen J. Characterization of type IV pilus genes in plant growth-promoting Pseudomonas putida WCS358. J Bacteriol. 1994 Feb;176(3):642–650. doi: 10.1128/jb.176.3.642-650.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]

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