Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Mar;161(3):1118–1124. doi: 10.1128/jb.161.3.1118-1124.1985

Protein export by a gram-negative bacterium: production of aerolysin by Aeromonas hydrophila.

S P Howard, J T Buckley
PMCID: PMC215015  PMID: 3972770

Abstract

The synthesis and export of aerolysin, an extracellular protein toxin released by the gram-negative bacterium Aeromonas hydrophila, was studied by pulse-labeling with [35S]methionine. The toxin was synthesized as a higher-molecular-weight precursor. This was processed cotranslationally, resulting in the appearance within the cell of the mature protein, which was then exported to the supernatant. Precursor aerolysin accumulated in cells incubated in the presence of carbonyl cyanide m-chlorophenyl hydrazone, a substance which also inhibited the export of mature aerolysin from the cell. The entrapped mature toxin could not be shocked from the cells, although it could be digested by protease applied to shocked cells. The toxin was processed and translocated across the inner membrane of pleiotropic export mutants and accumulated in the periplasm. The results indicate that more than one step is required for the export of the protein and that aerolysin does not cross the inner and outer membranes simultaneously.

Full text

PDF
1118

Images in this article

1119Image
on p.1119
1120Image
on p.1120
1120Image
on p.1120
1120Image
on p.1120
1121Image
on p.1121
1121Image
on p.1121
1122Image
on p.1122
1122Image
on p.1122

Selected References

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

  1. Anderson D. J., Blobel G. Immunoprecipitation of proteins from cell-free translations. Methods Enzymol. 1983;96:111–120. doi: 10.1016/s0076-6879(83)96012-3. [DOI] [PubMed] [Google Scholar]
  2. Enequist H. G., Hirst T. R., Harayama S., Hardy S. J., Randall L. L. Energy is required for maturation of exported proteins in Escherichia coli. Eur J Biochem. 1981 May 15;116(2):227–233. doi: 10.1111/j.1432-1033.1981.tb05323.x. [DOI] [PubMed] [Google Scholar]
  3. FINKELSTEIN R. A., NORRIS H. T., DUTTA N. K. PATHOGENESIS EXPERIMENTAL CHOLERA IN INFANT RABBITS. I. OBSERVATIONS ON THE INTRAINTESTINAL INFECTION AND EXPERIMENTAL CHOLERA PRODUCED WITH CELL-FREE PRODUCTS. J Infect Dis. 1964 Jun;114:203–216. doi: 10.1093/infdis/114.3.203. [DOI] [PubMed] [Google Scholar]
  4. Halpern Y. S., Lupo M. Glutamate transport in wild-type and mutant strains of Escherichia coli. J Bacteriol. 1965 Nov;90(5):1288–1295. doi: 10.1128/jb.90.5.1288-1295.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Hirst T. R., Randall L. L., Hardy S. J. Cellular location of heat-labile enterotoxin in Escherichia coli. J Bacteriol. 1984 Feb;157(2):637–642. doi: 10.1128/jb.157.2.637-642.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Howard S. P., Buckley J. T. Intracellular accumulation of extracellular proteins by pleiotropic export mutants of Aeromonas hydrophila. J Bacteriol. 1983 Apr;154(1):413–418. doi: 10.1128/jb.154.1.413-418.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Howard S. P., Buckley J. T. Membrane glycoprotein receptor and hole-forming properties of a cytolytic protein toxin. Biochemistry. 1982 Mar 30;21(7):1662–1667. doi: 10.1021/bi00536a029. [DOI] [PubMed] [Google Scholar]
  8. Ito K., Bassford P. J., Jr, Beckwith J. Protein localization in E. coli: is there a common step in the secretion of periplasmic and outer-membrane proteins? Cell. 1981 Jun;24(3):707–717. doi: 10.1016/0092-8674(81)90097-0. [DOI] [PubMed] [Google Scholar]
  9. Josefsson L. G., Randall L. L. Different exported proteins in E. coli show differences in the temporal mode of processing in vivo. Cell. 1981 Jul;25(1):151–157. doi: 10.1016/0092-8674(81)90239-7. [DOI] [PubMed] [Google Scholar]
  10. LEVY H. B., SOBER H. A. A simple chromatographic method for preparation of gamma globulin. Proc Soc Exp Biol Med. 1960 Jan;103:250–252. doi: 10.3181/00379727-103-25476. [DOI] [PubMed] [Google Scholar]
  11. Lopes J., Gottfried S., Rothfield L. Leakage of periplasmic enzymes by mutants of Escherichia coli and Salmonella typhimurium: isolation of "periplasmic leaky" mutants. J Bacteriol. 1972 Feb;109(2):520–525. doi: 10.1128/jb.109.2.520-525.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lory S., Tai P. C., Davis B. D. Mechanism of protein excretion by gram-negative bacteria: Pseudomonas aeruginosa exotoxin A. J Bacteriol. 1983 Nov;156(2):695–702. doi: 10.1128/jb.156.2.695-702.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Neill R. J., Ivins B. E., Holmes R. K. Synthesis and secretion of the plasmid-coded heat-labile enterotoxin of Escherichia coli in Vibrio cholerae. Science. 1983 Jul 15;221(4607):289–291. doi: 10.1126/science.6857285. [DOI] [PubMed] [Google Scholar]
  14. Neville D. M., Jr Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem. 1971 Oct 25;246(20):6328–6334. [PubMed] [Google Scholar]
  15. Pearson G. D., Mekalanos J. J. Molecular cloning of Vibrio cholerae enterotoxin genes in Escherichia coli K-12. Proc Natl Acad Sci U S A. 1982 May;79(9):2976–2980. doi: 10.1073/pnas.79.9.2976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Silhavy T. J., Benson S. A., Emr S. D. Mechanisms of protein localization. Microbiol Rev. 1983 Sep;47(3):313–344. doi: 10.1128/mr.47.3.313-344.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Wagner W., Vogel M., Goebel W. Transport of hemolysin across the outer membrane of Escherichia coli requires two functions. J Bacteriol. 1983 Apr;154(1):200–210. doi: 10.1128/jb.154.1.200-210.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Wretlind B., Pavlovskis O. R. Genetic mapping and characterization of Pseudomonas aeruginosa mutants defective in the formation of extracellular proteins. J Bacteriol. 1984 Jun;158(3):801–808. doi: 10.1128/jb.158.3.801-808.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Yamato I., Anraku Y., Hirosawa K. Cytoplasmic membrane vesicles of Escherichia coli. A simple method for preparing the cytoplasmic and outer membranes. J Biochem. 1975 Apr;77(4):705–718. doi: 10.1093/oxfordjournals.jbchem.a130774. [DOI] [PubMed] [Google Scholar]

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

RESOURCES