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. 1993 Dec;175(23):7639–7643. doi: 10.1128/jb.175.23.7639-7643.1993

Pap pili as a vector system for surface exposition of an immunoglobulin G-binding domain of protein A of Staphylococcus aureus in Escherichia coli.

L Steidler 1, E Remaut 1, W Fiers 1
PMCID: PMC206920  PMID: 7902348

Abstract

Fusion genes between papA, the gene coding for the major Pap pilus subunit, and fragments coding for an immunoglobulin G-binding domain of the Staphylococcus aureus protein A were constructed in such a way that the spa fragments were inserted following either codon 7 or 68 of the coding sequence for the mature portion of PapA. Peptides in the area of amino acids 7 and 68 of PapA are localized at the external side of the pilus. A set of pL expression plasmids containing papA and derivatives suitable for insertion were constructed. A papA gene carrying a spa insert following codon 68 was cloned back into the pap operon. The presence of this altered operon in a bacterial strain allowed the detection of immunoglobulin G-binding activity at the surfaces of the bacterial cells.

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Selected References

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  1. Abrahmsén L., Moks T., Nilsson B., Hellman U., Uhlén M. Analysis of signals for secretion in the staphylococcal protein A gene. EMBO J. 1985 Dec 30;4(13B):3901–3906. doi: 10.1002/j.1460-2075.1985.tb04164.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  4. Båga M., Normark S., Hardy J., O'Hanley P., Lark D., Olsson O., Schoolnik G., Falkow S. Nucleotide sequence of the papA gene encoding the Pap pilus subunit of human uropathogenic Escherichia coli. J Bacteriol. 1984 Jan;157(1):330–333. doi: 10.1128/jb.157.1.330-333.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Casadaban M. J., Cohen S. N. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. J Mol Biol. 1980 Apr;138(2):179–207. doi: 10.1016/0022-2836(80)90283-1. [DOI] [PubMed] [Google Scholar]
  6. Dougan G., Dowd G., Kehoe M. Organization of K88ac-encoded polypeptides in the Escherichia coli cell envelope: use of minicells and outer membrane protein mutants for studying assembly of pili. J Bacteriol. 1983 Jan;153(1):364–370. doi: 10.1128/jb.153.1.364-370.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Forsgren A., Sjöquist J. "Protein A" from S. aureus. I. Pseudo-immune reaction with human gamma-globulin. J Immunol. 1966 Dec;97(6):822–827. [PubMed] [Google Scholar]
  8. Hanson M. S., Hempel J., Brinton C. C., Jr Purification of the Escherichia coli type 1 pilin and minor pilus proteins and partial characterization of the adhesin protein. J Bacteriol. 1988 Aug;170(8):3350–3358. doi: 10.1128/jb.170.8.3350-3358.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hull R. A., Gill R. E., Hsu P., Minshew B. H., Falkow S. Construction and expression of recombinant plasmids encoding type 1 or D-mannose-resistant pili from a urinary tract infection Escherichia coli isolate. Infect Immun. 1981 Sep;33(3):933–938. doi: 10.1128/iai.33.3.933-938.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kahn M., Kolter R., Thomas C., Figurski D., Meyer R., Remaut E., Helinski D. R. Plasmid cloning vehicles derived from plasmids ColE1, F, R6K, and RK2. Methods Enzymol. 1979;68:268–280. doi: 10.1016/0076-6879(79)68019-9. [DOI] [PubMed] [Google Scholar]
  11. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  12. Lindberg F. P., Lund B., Normark S. Genes of pyelonephritogenic E. coli required for digalactoside-specific agglutination of human cells. EMBO J. 1984 May;3(5):1167–1173. doi: 10.1002/j.1460-2075.1984.tb01946.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lindberg F., Lund B., Johansson L., Normark S. Localization of the receptor-binding protein adhesin at the tip of the bacterial pilus. Nature. 1987 Jul 2;328(6125):84–87. doi: 10.1038/328084a0. [DOI] [PubMed] [Google Scholar]
  14. Lindberg F., Lund B., Normark S. Gene products specifying adhesion of uropathogenic Escherichia coli are minor components of pili. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1891–1895. doi: 10.1073/pnas.83.6.1891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Löfdahl S., Guss B., Uhlén M., Philipson L., Lindberg M. Gene for staphylococcal protein A. Proc Natl Acad Sci U S A. 1983 Feb;80(3):697–701. doi: 10.1073/pnas.80.3.697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Movitz J. Formation of extracellular protein A by Staphylococcus aureus. Eur J Biochem. 1976 Sep;68(1):291–299. doi: 10.1111/j.1432-1033.1976.tb10788.x. [DOI] [PubMed] [Google Scholar]
  17. Nakamaye K. L., Eckstein F. Inhibition of restriction endonuclease Nci I cleavage by phosphorothioate groups and its application to oligonucleotide-directed mutagenesis. Nucleic Acids Res. 1986 Dec 22;14(24):9679–9698. doi: 10.1093/nar/14.24.9679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Remaut E., Stanssens P., Fiers W. Inducible high level synthesis of mature human fibroblast interferon in Escherichia coli. Nucleic Acids Res. 1983 Jul 25;11(14):4677–4688. doi: 10.1093/nar/11.14.4677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Remaut E., Stanssens P., Fiers W. Plasmid vectors for high-efficiency expression controlled by the PL promoter of coliphage lambda. Gene. 1981 Oct;15(1):81–93. doi: 10.1016/0378-1119(81)90106-2. [DOI] [PubMed] [Google Scholar]
  20. Remaut E., Tsao H., Fiers W. Improved plasmid vectors with a thermoinducible expression and temperature-regulated runaway replication. Gene. 1983 Apr;22(1):103–113. doi: 10.1016/0378-1119(83)90069-0. [DOI] [PubMed] [Google Scholar]
  21. Schmidt M. A., O'Hanley P., Lark D., Schoolnik G. K. Synthetic peptides corresponding to protective epitopes of Escherichia coli digalactoside-binding pilin prevent infection in a murine pyelonephritis model. Proc Natl Acad Sci U S A. 1988 Feb;85(4):1247–1251. doi: 10.1073/pnas.85.4.1247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sjodahl J. Repetitive sequences in protein A from Staphylococcus aureus. Arrangement of five regions within the protein, four being highly homologous and Fc-binding. Eur J Biochem. 1977 Mar 1;73(2):343–351. doi: 10.1111/j.1432-1033.1977.tb11324.x. [DOI] [PubMed] [Google Scholar]
  23. Stanssens P., Opsomer C., McKeown Y. M., Kramer W., Zabeau M., Fritz H. J. Efficient oligonucleotide-directed construction of mutations in expression vectors by the gapped duplex DNA method using alternating selectable markers. Nucleic Acids Res. 1989 Jun 26;17(12):4441–4454. doi: 10.1093/nar/17.12.4441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Steidler L., Remaut E., Fiers W. LamB as a carrier molecule for the functional exposition of IgG-binding domains of the Staphylococcus aureus protein A at the surface of Escherichia coli K12. Mol Gen Genet. 1993 Jan;236(2-3):187–192. doi: 10.1007/BF00277111. [DOI] [PubMed] [Google Scholar]
  25. Uhlén M., Guss B., Nilsson B., Gatenbeck S., Philipson L., Lindberg M. Complete sequence of the staphylococcal gene encoding protein A. A gene evolved through multiple duplications. J Biol Chem. 1984 Feb 10;259(3):1695–1702. [PubMed] [Google Scholar]
  26. Van Die I., Wauben M., Van Megen I., Bergmans H., Riegman N., Hoekstra W., Pouwels P., Enger-Valk B. Genetic manipulation of major P-fimbrial subunits and consequences for formation of fimbriae. J Bacteriol. 1988 Dec;170(12):5870–5876. doi: 10.1128/jb.170.12.5870-5876.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Vogelstein B., Gillespie D. Preparative and analytical purification of DNA from agarose. Proc Natl Acad Sci U S A. 1979 Feb;76(2):615–619. doi: 10.1073/pnas.76.2.615. [DOI] [PMC free article] [PubMed] [Google Scholar]

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