Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1981 May;146(2):784–789. doi: 10.1128/jb.146.2.784-789.1981

Escherichia coli 987P pilus: purification and partial characterization.

R E Isaacson, P Richter
PMCID: PMC217025  PMID: 6111559

Abstract

The Escherichia coli somatic pilus, 987P, has been purified after removal by homogenization from a 987P+ enterotoxigenic E. coli. Cell-free pili were precipitated by the addition of MgCl2, collected, and dissolved in MgCl2-free buffer. Five cycles of precipitation and dissolving resulted in a preparation of 987P that was judged to be homogeneous based on electron microscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the electron microscope, 987P was rod shaped, having a diameter of 7 nm and an apparent axial hole. Cells and membrane vesicles were not observed in the purified pilus preparation. Electrophoresis of 987P through sodium dodecyl sulfate-polyacrylamide gels resulted in a single band when the sample was denatured in the absence of mercaptoethanol and in two bands when the sample was denatured in the presence of mercaptoethanol. The calculated molecular weight of 987 was variable, depending upon the polyacrylamide concentration and whether mercaptoethanol was included in the denaturing solution. Chemically, 987P is composed primarily of protein but also contains an unidentified amino sugar. The amino terminal amino acid of 987P is alanine and its isoelectric point is pH 3.7. 987P possesses no detectable hemagglutinating activity.

Full text

PDF
786

Images in this article

786Image
on p.786
787Image
on p.787

Selected References

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

  1. AMES B. N., DUBIN D. T. The role of polyamines in the neutralization of bacteriophage deoxyribonucleic acid. J Biol Chem. 1960 Mar;235:769–775. [PubMed] [Google Scholar]
  2. Brinton C. C., Jr The structure, function, synthesis and genetic control of bacterial pili and a molecular model for DNA and RNA transport in gram negative bacteria. Trans N Y Acad Sci. 1965 Jun;27(8):1003–1054. doi: 10.1111/j.2164-0947.1965.tb02342.x. [DOI] [PubMed] [Google Scholar]
  3. Edelhoch H. Spectroscopic determination of tryptophan and tyrosine in proteins. Biochemistry. 1967 Jul;6(7):1948–1954. doi: 10.1021/bi00859a010. [DOI] [PubMed] [Google Scholar]
  4. Fairbanks G., Steck T. L., Wallach D. F. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry. 1971 Jun 22;10(13):2606–2617. doi: 10.1021/bi00789a030. [DOI] [PubMed] [Google Scholar]
  5. Gros C., Labouesse B. Study of the dansylation reaction of amino acids, peptides and proteins. Eur J Biochem. 1969 Feb;7(4):463–470. doi: 10.1111/j.1432-1033.1969.tb19632.x. [DOI] [PubMed] [Google Scholar]
  6. HIRS C. H. The oxidation of ribonuclease with performic acid. J Biol Chem. 1956 Apr;219(2):611–621. [PubMed] [Google Scholar]
  7. Isaacson R. E., Dean E. A., Morgan R. L., Moon H. W. Immunization of suckling pigs against enterotoxigenic Escherichia coli-induced diarrheal disease by vaccinating dams with purified K99 or 987P pili: antibody production in response to vaccination. Infect Immun. 1980 Aug;29(2):824–826. doi: 10.1128/iai.29.2.824-826.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Isaacson R. E., Fusco P. C., Brinton C. C., Moon H. W. In vitro adhesion of Escherichia coli to porcine small intestinal epithelial cells: pili as adhesive factors. Infect Immun. 1978 Aug;21(2):392–397. doi: 10.1128/iai.21.2.392-397.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Isaacson R. E. K99 surface antigen of Escherichia coli: purification and partial characterization. Infect Immun. 1977 Jan;15(1):272–279. doi: 10.1128/iai.15.1.272-279.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Isaacson R. E., Nagy B., Moon H. W. Colonization of porcine small intestine by Escherichia coli: colonization and adhesion factors of pig enteropathogens that lack K88. J Infect Dis. 1977 Apr;135(4):531–539. doi: 10.1093/infdis/135.4.531. [DOI] [PubMed] [Google Scholar]
  11. Jones G. W., Rutter J. M. The association of K88 antigen with haemagglutinating activity in porcine strains of Escherichia coli. J Gen Microbiol. 1974 Sep;84(1):135–144. doi: 10.1099/00221287-84-1-135. [DOI] [PubMed] [Google Scholar]
  12. Klemm P. Fimbrial colonization factor CFA/1 protein from human enteropathogenic Escherichia coli strains. Purification, characterization and N-terminal sequence. FEBS Lett. 1979 Dec 1;108(1):107–110. doi: 10.1016/0014-5793(79)81188-6. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. McMichael J. C., Ou J. T. Structure of common pili from Escherichia coli. J Bacteriol. 1979 Jun;138(3):969–975. doi: 10.1128/jb.138.3.969-975.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Moon H. W., Kohler E. M., Schneider R. A., Whipp S. C. Prevalence of pilus antigens, enterotoxin types, and enteropathogenicity among K88-negative enterotoxigenic Escherichia coli from neonatal pigs. Infect Immun. 1980 Jan;27(1):222–230. doi: 10.1128/iai.27.1.222-230.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Morgan R. L., Isaacson R. E., Moon H. W., Brinton C. C., To C. C. Immunization of suckling pigs against enterotoxigenic Escherichia coli-induced diarrheal disease by vaccinating dams with purified 987 or K99 pili: protection correlates with pilus homology of vaccine and challenge. Infect Immun. 1978 Dec;22(3):771–777. doi: 10.1128/iai.22.3.771-777.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nagy B., Moon H. W., Isaacson R. E. Colonization of porcine intestine by enterotoxigenic Escherichia coli: selection of piliated forms in vivo, adhesion of piliated forms to epithelial cells in vitro, and incidence of a pilus antigen among porcine enteropathogenic E. coli. Infect Immun. 1977 Apr;16(1):344–352. doi: 10.1128/iai.16.1.344-352.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nagy B., Moon H. W., Isaacson R. E. Colonization of porcine small intestine by Escherichia coli: ileal colonization and adhesion by pig enteropathogens that lack K88 antigen and by some acapsular mutants. Infect Immun. 1976 Apr;13(4):1214–1220. doi: 10.1128/iai.13.4.1214-1220.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Nagy B., Moon H. W., Isaacson R. E., To C. C., Brinton C. C. Immunization of suckling pigs against enteric enterotoxigenic Escherichia coli infection by vaccinating dams with purified pili. Infect Immun. 1978 Jul;21(1):269–274. doi: 10.1128/iai.21.1.269-274.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Salit I. E., Gotschlich E. C. Hemagglutination by purified type I Escherichia coli pili. J Exp Med. 1977 Nov 1;146(5):1169–1181. doi: 10.1084/jem.146.5.1169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Smith H. W., Linggood M. A. Further observations on Escherichia coli enterotoxins with particular regard to those produced by atypical piglet strains and by calf and lamb strains: the transmissible nature of these enterotoxins and of a K antigen possessed by calf and lamb strains. J Med Microbiol. 1972 May;5(2):243–250. doi: 10.1099/00222615-5-2-243. [DOI] [PubMed] [Google Scholar]
  23. Smith H. W., Linggood M. A. Observations on the pathogenic properties of the K88, Hly and Ent plasmids of Escherichia coli with particular reference to porcine diarrhoea. J Med Microbiol. 1971 Nov;4(4):467–485. doi: 10.1099/00222615-4-4-467. [DOI] [PubMed] [Google Scholar]
  24. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]

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

RESOURCES