Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1992 Apr;60(4):1302–1313. doi: 10.1128/iai.60.4.1302-1313.1992

Cloning, expression, and DNA sequence analysis of genes encoding nontypeable Haemophilus influenzae high-molecular-weight surface-exposed proteins related to filamentous hemagglutinin of Bordetella pertussis.

S J Barenkamp 1, E Leininger 1
PMCID: PMC256997  PMID: 1548058

Abstract

A group of high-molecular-weight surface-exposed proteins of nontypeable Haemophilus influenzae are major targets of human serum antibody (S. J. Barenkamp and F. F. Bodor, Pediatr. Infect. Dis. J. 9:333-337, 1990). To further characterize these proteins, we cloned and sequenced genes encoding two related high-molecular-weight proteins from a prototype nontypeable Haemophilus strain. The gene encoding a 120-kDa Haemophilus protein consisted of a 4.4-kbp open reading frame, and the gene encoding a 125-kDa protein consisted of a 4.6-kbp open reading frame. The first 1,259 bp of the two genes were identical. Thereafter, the sequences began to diverge, but overall they were 80% identical, and the derived amino acid sequences showed 70% identity. A protein sequence homology search demonstrated similarity between the derived amino acid sequences of both cloned genes and the derived amino acid sequence of the gene encoding filamentous hemagglutinin, a surface protein produced by the gram-negative pathogen Bordetella pertussis. Antiserum raised against a recombinant protein encoded by the 4.6-kbp open reading frame recognized both the 120- and the 125-kDa proteins in the prototype strain as well as antigenically related high-molecular-weight proteins in 75% of a collection of 125 epidemiologically unrelated nontypeable H. influenzae strains. The antiserum directed against the recombinant protein also recognized purified filamentous hemagglutinin. A murine monoclonal antibody to filamentous hemagglutinin recognized both the 120-kDa and the 125-kDa protein in the prototype strain as well as proteins identical to those recognized by the recombinant-protein antiserum in 35% of the nontypeable H. influenzae strain collection. Thus, we have identified and partially characterized a group of highly immunogenic surface-exposed proteins of nontypeable H. influenzae which are related to the filamentous hemagglutinin of B. pertussis.

Full text

PDF
1305

Images in this article

1304Image
on p.1304
1306Image
on p.1306
1311Image
on p.1311
1311Image
on p.1311

Selected References

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

  1. Barenkamp S. J., Bodor F. F. Development of serum bactericidal activity following nontypable Haemophilus influenzae acute otitis media. Pediatr Infect Dis J. 1990 May;9(5):333–339. doi: 10.1097/00006454-199005000-00006. [DOI] [PubMed] [Google Scholar]
  2. Barnes W. M. Sequencing DNA with dideoxyribonucleotides as chain terminators: hints and strategies for big projects. Methods Enzymol. 1987;152:538–556. doi: 10.1016/0076-6879(87)52060-2. [DOI] [PubMed] [Google Scholar]
  3. Brendel V., Trifonov E. N. A computer algorithm for testing potential prokaryotic terminators. Nucleic Acids Res. 1984 May 25;12(10):4411–4427. doi: 10.1093/nar/12.10.4411. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Delisse-Gathoye A. M., Locht C., Jacob F., Raaschou-Nielsen M., Heron I., Ruelle J. L., de Wilde M., Cabezon T. Cloning, partial sequence, expression, and antigenic analysis of the filamentous hemagglutinin gene of Bordetella pertussis. Infect Immun. 1990 Sep;58(9):2895–2905. doi: 10.1128/iai.58.9.2895-2905.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Domenighini M., Relman D., Capiau C., Falkow S., Prugnola A., Scarlato V., Rappuoli R. Genetic characterization of Bordetella pertussis filamentous haemagglutinin: a protein processed from an unusually large precursor. Mol Microbiol. 1990 May;4(5):787–800. doi: 10.1111/j.1365-2958.1990.tb00649.x. [DOI] [PubMed] [Google Scholar]
  7. Ely S., Tippett J., Kroll J. S., Moxon E. R. Mutations affecting expression and maintenance of genes encoding the serotype b capsule of Haemophilus influenzae. J Bacteriol. 1986 Jul;167(1):44–48. doi: 10.1128/jb.167.1.44-48.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Farley M. M., Stephens D. S., Kaplan S. L., Mason E. O., Jr Pilus- and non-pilus-mediated interactions of Haemophilus influenzae type b with human erythrocytes and human nasopharyngeal mucosa. J Infect Dis. 1990 Feb;161(2):274–280. doi: 10.1093/infdis/161.2.274. [DOI] [PubMed] [Google Scholar]
  9. Gnehm H. E., Pelton S. I., Gulati S., Rice P. A. Characterization of antigens from nontypable Haemophilus influenzae recognized by human bactericidal antibodies. Role of Haemophilus outer membrane proteins. J Clin Invest. 1985 May;75(5):1645–1658. doi: 10.1172/JCI111872. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hallander H. O., Storsaeter J., Möllby R. Evaluation of serology and nasopharyngeal cultures for diagnosis of pertussis in a vaccine efficacy trial. J Infect Dis. 1991 May;163(5):1046–1054. doi: 10.1093/infdis/163.5.1046. [DOI] [PubMed] [Google Scholar]
  11. Kimura A., Mountzouros K. T., Relman D. A., Falkow S., Cowell J. L. Bordetella pertussis filamentous hemagglutinin: evaluation as a protective antigen and colonization factor in a mouse respiratory infection model. Infect Immun. 1990 Jan;58(1):7–16. doi: 10.1128/iai.58.1.7-16.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. 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]
  13. Loeb M. R., Connor E., Penney D. A comparison of the adherence of fimbriated and nonfimbriated Haemophilus influenzae type b to human adenoids in organ culture. Infect Immun. 1988 Feb;56(2):484–489. doi: 10.1128/iai.56.2.484-489.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. Miller V. L., Mekalanos J. J. Synthesis of cholera toxin is positively regulated at the transcriptional level by toxR. Proc Natl Acad Sci U S A. 1984 Jun;81(11):3471–3475. doi: 10.1073/pnas.81.11.3471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Miller V. L., Taylor R. K., Mekalanos J. J. Cholera toxin transcriptional activator toxR is a transmembrane DNA binding protein. Cell. 1987 Jan 30;48(2):271–279. doi: 10.1016/0092-8674(87)90430-2. [DOI] [PubMed] [Google Scholar]
  17. Moffatt B. A., Studier F. W. T7 lysozyme inhibits transcription by T7 RNA polymerase. Cell. 1987 Apr 24;49(2):221–227. doi: 10.1016/0092-8674(87)90563-0. [DOI] [PubMed] [Google Scholar]
  18. Murphy T. F., Apicella M. A. Nontypable Haemophilus influenzae: a review of clinical aspects, surface antigens, and the human immune response to infection. Rev Infect Dis. 1987 Jan-Feb;9(1):1–15. doi: 10.1093/clinids/9.1.1. [DOI] [PubMed] [Google Scholar]
  19. Murphy T. F., Bartos L. C. Human bactericidal antibody response to outer membrane protein P2 of nontypeable Haemophilus influenzae. Infect Immun. 1988 Oct;56(10):2673–2679. doi: 10.1128/iai.56.10.2673-2679.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Murphy T. F., Bartos L. C., Rice P. A., Nelson M. B., Dudas K. C., Apicella M. A. Identification of a 16,600-dalton outer membrane protein on nontypeable Haemophilus influenzae as a target for human serum bactericidal antibody. J Clin Invest. 1986 Oct;78(4):1020–1027. doi: 10.1172/JCI112656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Musser J. M., Barenkamp S. J., Granoff D. M., Selander R. K. Genetic relationships of serologically nontypable and serotype b strains of Haemophilus influenzae. Infect Immun. 1986 Apr;52(1):183–191. doi: 10.1128/iai.52.1.183-191.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Oda M., Cowell J. L., Burstyn D. G., Manclark C. R. Protective activities of the filamentous hemagglutinin and the lymphocytosis-promoting factor of Bordetella pertussis in mice. J Infect Dis. 1984 Dec;150(6):823–833. doi: 10.1093/infdis/150.6.823. [DOI] [PubMed] [Google Scholar]
  23. Poole K., Schiebel E., Braun V. Molecular characterization of the hemolysin determinant of Serratia marcescens. J Bacteriol. 1988 Jul;170(7):3177–3188. doi: 10.1128/jb.170.7.3177-3188.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Read R. C., Wilson R., Rutman A., Lund V., Todd H. C., Brain A. P., Jeffery P. K., Cole P. J. Interaction of nontypable Haemophilus influenzae with human respiratory mucosa in vitro. J Infect Dis. 1991 Mar;163(3):549–558. doi: 10.1093/infdis/163.3.549. [DOI] [PubMed] [Google Scholar]
  25. Relman D. A., Domenighini M., Tuomanen E., Rappuoli R., Falkow S. Filamentous hemagglutinin of Bordetella pertussis: nucleotide sequence and crucial role in adherence. Proc Natl Acad Sci U S A. 1989 Apr;86(8):2637–2641. doi: 10.1073/pnas.86.8.2637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Relman D., Tuomanen E., Falkow S., Golenbock D. T., Saukkonen K., Wright S. D. Recognition of a bacterial adhesion by an integrin: macrophage CR3 (alpha M beta 2, CD11b/CD18) binds filamentous hemagglutinin of Bordetella pertussis. Cell. 1990 Jun 29;61(7):1375–1382. doi: 10.1016/0092-8674(90)90701-f. [DOI] [PubMed] [Google Scholar]
  27. Sable N. S., Connor E. M., Hall C. B., Loeb M. R. Variable adherence of fimbriated Haemophilus influenzae type b to human cells. Infect Immun. 1985 Apr;48(1):119–123. doi: 10.1128/iai.48.1.119-123.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Smith P. K., Krohn R. I., Hermanson G. T., Mallia A. K., Gartner F. H., Provenzano M. D., Fujimoto E. K., Goeke N. M., Olson B. J., Klenk D. C. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985 Oct;150(1):76–85. doi: 10.1016/0003-2697(85)90442-7. [DOI] [PubMed] [Google Scholar]
  29. St Geme J. W., 3rd, Falkow S. Haemophilus influenzae adheres to and enters cultured human epithelial cells. Infect Immun. 1990 Dec;58(12):4036–4044. doi: 10.1128/iai.58.12.4036-4044.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Stibitz S., Weiss A. A., Falkow S. Genetic analysis of a region of the Bordetella pertussis chromosome encoding filamentous hemagglutinin and the pleiotropic regulatory locus vir. J Bacteriol. 1988 Jul;170(7):2904–2913. doi: 10.1128/jb.170.7.2904-2913.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. 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]
  32. Tabor S., Richardson C. C. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. doi: 10.1073/pnas.82.4.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Tuomanen E., Towbin H., Rosenfelder G., Braun D., Larson G., Hansson G. C., Hill R. Receptor analogs and monoclonal antibodies that inhibit adherence of Bordetella pertussis to human ciliated respiratory epithelial cells. J Exp Med. 1988 Jul 1;168(1):267–277. doi: 10.1084/jem.168.1.267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tuomanen E., Weiss A. Characterization of two adhesins of Bordetella pertussis for human ciliated respiratory-epithelial cells. J Infect Dis. 1985 Jul;152(1):118–125. doi: 10.1093/infdis/152.1.118. [DOI] [PubMed] [Google Scholar]
  35. Uphoff T. S., Welch R. A. Nucleotide sequencing of the Proteus mirabilis calcium-independent hemolysin genes (hpmA and hpmB) reveals sequence similarity with the Serratia marcescens hemolysin genes (shlA and shlB). J Bacteriol. 1990 Mar;172(3):1206–1216. doi: 10.1128/jb.172.3.1206-1216.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Urisu A., Cowell J. L., Manclark C. R. Filamentous hemagglutinin has a major role in mediating adherence of Bordetella pertussis to human WiDr cells. Infect Immun. 1986 Jun;52(3):695–701. doi: 10.1128/iai.52.3.695-701.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wallace R. J., Jr, Musher D. M., Septimus E. J., McGowan J. E., Jr, Quinones F. J., Wiss K., Vance P. H., Trier P. A. Haemophilus influenzae infections in adults: characterization of strains by serotypes, biotypes, and beta-lactamase production. J Infect Dis. 1981 Aug;144(2):101–106. doi: 10.1093/infdis/144.2.101. [DOI] [PubMed] [Google Scholar]

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

RESOURCES