Skip to main content
Infection and Immunity logoLink to Infection and Immunity
. 1989 Nov;57(11):3663–3665. doi: 10.1128/iai.57.11.3663-3665.1989

Fusion of genes encoding Escherichia coli heat-stable enterotoxin and outer membrane protein OmpC.

H T Saarilahti 1, E T Palva 1, J Holmgren 1, J Sanchez 1
PMCID: PMC259882  PMID: 2680976

Abstract

The OmpC outer membrane protein of Escherichia coli was used as a carrier molecule for the nonimmunogenic heat-stable enterotoxin STa. Two fragments of different lengths of the gene encoding STa were fused in vitro to the 3' terminus of the truncated ompC gene. The resulting OmpC-STa hybrid proteins could be detected by L-[35S]cysteine labeling, and they were processed and thus exported. All synthesized hybrid protein remained cell bound and was found by fractionation mainly in the periplasm. Immunoblot analysis showed that the hybrid proteins reacted in vitro both with anti-OmpC and anti-STa antibodies, and immunization of rabbits evoked an antibody response to either of these proteins.

Full text

PDF
3663

Images in this article

Selected References

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

  1. Burgess M. N., Bywater R. J., Cowley C. M., Mullan N. A., Newsome P. M. Biological evaluation of a methanol-soluble, heat-stable Escherichia coli enterotoxin in infant mice, pigs, rabbits, and calves. Infect Immun. 1978 Aug;21(2):526–531. doi: 10.1128/iai.21.2.526-531.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  3. Field M., Graf L. H., Jr, Laird W. J., Smith P. L. Heat-stable enterotoxin of Escherichia coli: in vitro effects on guanylate cyclase activity, cyclic GMP concentration, and ion transport in small intestine. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2800–2804. doi: 10.1073/pnas.75.6.2800. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Guzman-Verduzco L. M., Kupersztoch Y. M. Fusion of Escherichia coli heat-stable enterotoxin and heat-labile enterotoxin B subunit. J Bacteriol. 1987 Nov;169(11):5201–5208. doi: 10.1128/jb.169.11.5201-5208.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Klipstein F. A., Engert R. F., Clements J. D., Houghten R. A. Vaccine for enterotoxigenic Escherichia coli based on synthetic heat-stable toxin crossed-linked to the B subunit of heat-labile toxin. J Infect Dis. 1983 Feb;147(2):318–326. doi: 10.1093/infdis/147.2.318. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. Lee C. H., Moseley S. L., Moon H. W., Whipp S. C., Gyles C. L., So M. Characterization of the gene encoding heat-stable toxin II and preliminary molecular epidemiological studies of enterotoxigenic Escherichia coli heat-stable toxin II producers. Infect Immun. 1983 Oct;42(1):264–268. doi: 10.1128/iai.42.1.264-268.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Liljeström P., Mättänen P. L., Palva E. T. Cloning of the regulatory locus ompB of Salmonella typhimurium LT-2. II. Identification of the envZ gene product, a protein involved in the expression of the porin proteins. Mol Gen Genet. 1982;188(2):190–194. doi: 10.1007/BF00332674. [DOI] [PubMed] [Google Scholar]
  9. Mizuno T., Chou M. Y., Inouye M. A comparative study on the genes for three porins of the Escherichia coli outer membrane. DNA sequence of the osmoregulated ompC gene. J Biol Chem. 1983 Jun 10;258(11):6932–6940. [PubMed] [Google Scholar]
  10. Nikaido H., Vaara M. Molecular basis of bacterial outer membrane permeability. Microbiol Rev. 1985 Mar;49(1):1–32. doi: 10.1128/mr.49.1.1-32.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  12. Osborn M. J., Gander J. E., Parisi E., Carson J. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Isolation and characterization of cytoplasmic and outer membrane. J Biol Chem. 1972 Jun 25;247(12):3962–3972. [PubMed] [Google Scholar]
  13. Palva E. T., Hirst T. R., Hardy S. J., Holmgren J., Randall L. Synthesis of a precursor to the B subunit of heat-labile enterotoxin in Escherichia coli. J Bacteriol. 1981 Apr;146(1):325–330. doi: 10.1128/jb.146.1.325-330.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Sack R. B. Human diarrheal disease caused by enterotoxigenic Escherichia coli. Annu Rev Microbiol. 1975;29:333–353. doi: 10.1146/annurev.mi.29.100175.002001. [DOI] [PubMed] [Google Scholar]
  15. Sancar A., Hack A. M., Rupp W. D. Simple method for identification of plasmid-coded proteins. J Bacteriol. 1979 Jan;137(1):692–693. doi: 10.1128/jb.137.1.692-693.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sanchez J., Hirst T. R., Uhlin B. E. Hybrid enterotoxin LTA::STa proteins and their protection from degradation by in vivo association with B-subunits of Escherichia coli heat-labile enterotoxin. Gene. 1988 Apr 29;64(2):265–275. doi: 10.1016/0378-1119(88)90341-1. [DOI] [PubMed] [Google Scholar]
  17. Sanchez J., Svennerholm A. M., Holmgren J. Genetic fusion of a non-toxic heat-stable enterotoxin-related decapeptide antigen to cholera toxin B-subunit. FEBS Lett. 1988 Dec 5;241(1-2):110–114. doi: 10.1016/0014-5793(88)81041-x. [DOI] [PubMed] [Google Scholar]
  18. Sanchez J., Uhlin B. E., Grundström T., Holmgren J., Hirst T. R. Immunoactive chimeric ST-LT enterotoxins of Escherichia coli generated by in vitro gene fusion. FEBS Lett. 1986 Nov 24;208(2):194–198. doi: 10.1016/0014-5793(86)81016-x. [DOI] [PubMed] [Google Scholar]
  19. So M., McCarthy B. J. Nucleotide sequence of the bacterial transposon Tn1681 encoding a heat-stable (ST) toxin and its identification in enterotoxigenic Escherichia coli strains. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4011–4015. doi: 10.1073/pnas.77.7.4011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Svennerholm A. M., Wikström M., Lindblad M., Holmgren J. Monoclonal antibodies against Escherichia coli heat-stable toxin (STa) and their use in a diagnostic ST ganglioside GM1-enzyme-linked immunosorbent assay. J Clin Microbiol. 1986 Oct;24(4):585–590. doi: 10.1128/jcm.24.4.585-590.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES