Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Jun;162(3):1075–1078. doi: 10.1128/jb.162.3.1075-1078.1985

Expression of Actinomyces viscosus antigens in Escherichia coli: cloning of a structural gene (fimA) for type 2 fimbriae.

J A Donkersloot, J O Cisar, M E Wax, R J Harr, B M Chassy
PMCID: PMC215885  PMID: 2860095

Abstract

A cosmid gene library of Actinomyces viscosus T14V was prepared in Escherichia coli to examine the expression of A. viscosus antigens and to gain insight into the structure of A. viscosus type 1 and type 2 fimbriae. Out of this library of 550 clones, 28 reacted in a colony immunoassay with antibodies against A. viscosus cells. The proteins responsible for these reactions were identified in three clones. Clones AV1209 and AV2009 displayed nonfimbrial antigens with subunits of 40 and 58 kilodaltons, respectively. Clone AV1402 showed a 59-kilodalton protein that reacted with monospecific antibody against type 2 fimbriae and that comigrated with a subunit of type 2 fimbriae during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This indicates that AV1402 expresses a gene (fimA) for a subunit of A. viscosus type 2 fimbriae.

Full text

PDF
1078

Images in this article

1076Image
on p.1076
1077Image
on p.1077
1077Image
on p.1077

Selected References

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

  1. Batteiger B., Newhall W. J., 5th, Jones R. B. The use of Tween 20 as a blocking agent in the immunological detection of proteins transferred to nitrocellulose membranes. J Immunol Methods. 1982 Dec 30;55(3):297–307. doi: 10.1016/0022-1759(82)90089-8. [DOI] [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. Brammar W. J., Muir S., McMorris A. Molecular cloning of the gene for the beta-lactamase of Bacillus licheniformis and its expression in Escherichia coli. Mol Gen Genet. 1980 Apr;178(1):217–224. doi: 10.1007/BF00267232. [DOI] [PubMed] [Google Scholar]
  5. Chassy B. M., Giuffrida A. Method for the lysis of Gram-positive, asporogenous bacteria with lysozyme. Appl Environ Microbiol. 1980 Jan;39(1):153–158. doi: 10.1128/aem.39.1.153-158.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cisar J. O., Barsumian E. L., Curl S. H., Vatter A. E., Sandberg A. L., Siraganian R. P. Detection and localization of a lectin on Actinomyces viscosus T14V by monoclonal antibodies. J Immunol. 1981 Oct;127(4):1318–1322. [PubMed] [Google Scholar]
  7. Cisar J. O., Curl S. H., Kolenbrander P. E., Vatter A. E. Specific absence of type 2 fimbriae on a coaggregation-defective mutant of Actinomyces viscosus T14V. Infect Immun. 1983 May;40(2):759–765. doi: 10.1128/iai.40.2.759-765.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clark W. B., Wheeler T. T., Cisar J. O. Specific inhibition of adsorption of Actinomyces viscosus T14V to saliva-treated hydroxyapatite by antibody against type 1 fimbriae. Infect Immun. 1984 Feb;43(2):497–501. doi: 10.1128/iai.43.2.497-501.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  10. Cornelis P., Digneffe C., Willemot K. Cloning and expression of a Bacillus coagulans amylase gene in Escherichia coli. Mol Gen Genet. 1982;186(4):507–511. doi: 10.1007/BF00337957. [DOI] [PubMed] [Google Scholar]
  11. Cuatrecasas P. Protein purification by affinity chromatography. Derivatizations of agarose and polyacrylamide beads. J Biol Chem. 1970 Jun;245(12):3059–3065. [PubMed] [Google Scholar]
  12. Ehrlich S. D., Bursztyn-Pettegrew H., Stroynowski I., Lederberg J. Expression of the thymidylate synthetase gene of the Bacillus subtilis bacteriophage Phi-3-T in Escherichia coli. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4145–4149. doi: 10.1073/pnas.73.11.4145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Guerry P., LeBlanc D. J., Falkow S. General method for the isolation of plasmid deoxyribonucleic acid. J Bacteriol. 1973 Nov;116(2):1064–1066. doi: 10.1128/jb.116.2.1064-1066.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Helfman D. M., Feramisco J. R., Fiddes J. C., Thomas G. P., Hughes S. H. Identification of clones that encode chicken tropomyosin by direct immunological screening of a cDNA expression library. Proc Natl Acad Sci U S A. 1983 Jan;80(1):31–35. doi: 10.1073/pnas.80.1.31. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hohn B., Collins J. A small cosmid for efficient cloning of large DNA fragments. Gene. 1980 Nov;11(3-4):291–298. doi: 10.1016/0378-1119(80)90069-4. [DOI] [PubMed] [Google Scholar]
  16. Holt R. G., Abiko Y., Saito S., Smorawinska M., Hansen J. B., Curtiss R., 3rd Streptococcus mutans genes that code for extracellular proteins in Escherichia coli K-12. Infect Immun. 1982 Oct;38(1):147–156. doi: 10.1128/iai.38.1.147-156.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jagusztyn-Krynicka E. K., Smorawinska M., Curtiss R., 3rd Expression of Streptococcus mutans aspartate-semialdehyde dehydrogenase gene cloned into plasmid pBR322. J Gen Microbiol. 1982 May;128(5):1135–1145. doi: 10.1099/00221287-128-5-1135. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Lee L. J., Hansen J. B., Jagusztyn-Krynicka E. K., Chassy B. M. Cloning and expression of the beta-D-phosphogalactoside galactohydrolase gene of Lactobacillus casei in Escherichia coli K-12. J Bacteriol. 1982 Dec;152(3):1138–1146. doi: 10.1128/jb.152.3.1138-1146.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Masuda N., Ellen R. P., Fillery E. D., Grove D. A. Chemical and immunological comparison of surface fibrils of strains representing six taxonomic groups of Actinomyces viscosus and Actinomyces naeslundii. Infect Immun. 1983 Mar;39(3):1325–1333. doi: 10.1128/iai.39.3.1325-1333.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. O'Farrell P. H., Kutter E., Nakanishi M. A restriction map of the bacteriophage T4 genome. Mol Gen Genet. 1980;179(2):421–435. doi: 10.1007/BF00425473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Revis G. J., Vatter A. E., Crowle A. J., Cisar J. O. Antibodies against the Ag2 fimbriae of Actinomyces viscosus T14V inhibit lactose-sensitive bacterial adherence. Infect Immun. 1982 Jun;36(3):1217–1222. doi: 10.1128/iai.36.3.1217-1222.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Scott J. R., Fischetti V. A. Expression of streptococcal M protein in Escherichia coli. Science. 1983 Aug 19;221(4612):758–760. doi: 10.1126/science.6192499. [DOI] [PubMed] [Google Scholar]
  24. Stüber D., Bujard H. Organization of transcriptional signals in plasmids pBR322 and pACYC184. Proc Natl Acad Sci U S A. 1981 Jan;78(1):167–171. doi: 10.1073/pnas.78.1.167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wheeler T. T., Clark W. B. Fibril-mediated adherence of Actinomyces viscosus to saliva-treated hydroxyapatite. Infect Immun. 1980 May;28(2):577–584. doi: 10.1128/iai.28.2.577-584.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES