1887

Abstract

The gene cluster of subsp. (Ecc) encodes the proteins of the type II or general secretory pathway (GSP) apparatus which is required for secretion of pectinase and cellulase. In this study, fusions between Ecc genes and the topology probe were constructed. The ability of Out protein domains to export BlaM across the cytoplasmic membrane in both and the cognate host was utilized to confirm the computer-predicted cytoplasmic membrane topology of OutC and OutF. When was fused to , the resulting phenotype suggested that the majority of OutC is targeted to the periplasm, typical of a type II bitopic conformation in the cytoplasmic membrane. In contrast, for the gene product, three transmembrane regions were identified which connect a large N-terminal cytoplasmic domain, a smaller periplasmic domain, and a large cytoplasmic loop. Fusions between and and were used to further substantiate the locations of these gene products in the outer membrane and the cytoplasm respectively. The data derived suggest that a number of the Out apparatus components possess domains in the cytoplasm and/or the periplasm with potential for protein-protein interactions which facilitate the secretion of periplasmic enzyme intermediates across the outer membrane to the external milieu.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-143-3-713
1997-03-01
2021-05-06
Loading full text...

Full text loading...

/deliver/fulltext/micro/143/3/mic-143-3-713.html?itemId=/content/journal/micro/10.1099/00221287-143-3-713&mimeType=html&fmt=ahah

References

  1. Bleves S., Lazdunski A., Filloux A. 1996; Membrane topology of three Xcp proteins involved in exoprotein transport by Pseudomonas aeruginosa.. J Bacteriol 178:4297–4300
    [Google Scholar]
  2. Broome-Smith J.K., Tadayyon M., Zhang Y. 1990; β-Lactamase as a probe of membrane protein assembly and protein export.. Mol Microbiol 4:1637–1644
    [Google Scholar]
  3. Clement J.M., Perrin D., Hedgpeth J. 1982; Analysis of a receptor and ^-lactamase synthesis and export using cloned genes in a minicell system.. Mol Gen Genet 185:302–310
    [Google Scholar]
  4. d´Enfert C., Reyss I., Wandersman C., Pugsley A.P. 1989; Protein secretion by Gram-negative bacteria. Characterization of two membrane proteins required for pullulanase secretion by Escherichia coli K-12.. J Biol Chem 264:17462–17468
    [Google Scholar]
  5. Edelman A., Bowler L., Broome-Smith J.K., Spratt B.G. 1987; Use of a β-lactamase fusion vector to investigate the organization of penicillin-binding protein 1B in the cytoplasmic membrane of Escherichia coli.. Mol Microbiol 1:101–106
    [Google Scholar]
  6. Gibson T.J. 1984 Studies on Epstein-Barr virus genome. PhD thesis, Cambridge University, England
    [Google Scholar]
  7. Hanahan D. 1983; Studies on transformation of Escherichia coli with plasmids.. J Mol Biol 166:557–580
    [Google Scholar]
  8. Hardie K.R., Lory S., Pugsley A.P. 1996; Insertion of an outer membrane protein in Escherichia coli requires a chaperone-like protein.. EMBO J 15:978–988
    [Google Scholar]
  9. Hu N., Hung M., Liao C., Lin M. 1995; Subcellular location of XpsD, a protein required for extracellular protein secretion by Xanthomonas campestris pv. campestris.. Microbiology 141:1395–1406
    [Google Scholar]
  10. Jaurin B., Grundström T., Normark S. 1982; Sequence elements determining ampC promoter strength in Escherichia coli.. EMBO J 1:875–881
    [Google Scholar]
  11. Kazmierczak B.I., Mielke D.L., Russel M., Model P. 1994; pIV, a filamentous phage protein that mediates phage export across the bacterial cell envelope, forms a multimer.. J Mol Biol 238:187–198
    [Google Scholar]
  12. Kyte J., Doolittle R.F. 1982; A simple method for displaying the hydrophobic character of a protein.. J Mol Biol 157:105–132
    [Google Scholar]
  13. Lindeberg M., Salmond G.P.C., Collmer A. 1996; Complementation of deletion mutations in a cloned functional cluster of Erwinia chrysanthemi out genes with Erwinia carotovora out homologues reveals OutC and OutD as candidate gatekeepers of species-specific secretion of proteins via the type II pathway.. Mol Microbiol 20:175–190
    [Google Scholar]
  14. Miller J.H. 1972 Experiments in Molecular Genetics. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  15. Pugsley A. 1993; The complete general secretory pathway in Gram-negative bacteria.. Microbiol Rev 57:50–108
    [Google Scholar]
  16. Queen C., Korn L.J. 1984; A comprehensive sequence analysis programme for the IBM personal computer.. Nucleic Acids Res 12:581–599
    [Google Scholar]
  17. Reeves P.J., Whitcombe D., Wharam S., Gibson M., Allison G., Bunce N., Barallon R., Douglas P., Mulholland V., Stevens S., Walker D., Salmond G.P.C. 1993; Molecular cloning and characterization of 13 out genes from Erwinia carotovora subspecies carotovora-. genes encoding members of a general secretion pathway (GSP) widespread in Gram-negative bacteria.. Mol Microbiol 8:433–456
    [Google Scholar]
  18. Reeves P.J., Douglas P., Salmond G.P.C. 1994; The general secretion pathway of Erwinia carotovora subspecies carotovora: analysis of the cytoplasmic membrane topology of seven Out proteins using a β-lactamase topology probe.. Mol Microbiol 12:445–457
    [Google Scholar]
  19. Russel M., Kazmierczak B. 1993; Analysis of the structure and subcellular location of filamentous phage pIV.. J Bacteriol 175:3998–4007
    [Google Scholar]
  20. Salmond G.P.C., Reeves P.J. 1993; Membrane traffic wardens and protein secretion in Gram-negative bacteria.. Trends Biochem Sci 18:7–12
    [Google Scholar]
  21. Sambrook J., Fritsch E.F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd edn.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  22. Sandkvist M., Bagdasarian M., Howard S.P., Dirita V.J. 1995; Interaction between the autokinase EpsE and EpsL in the cytoplasmic membrane is required for extracellular secretion in Vibrio cholerae.. EMBO J 14:1664–1673
    [Google Scholar]
  23. Wharam S.D., Mulholland V., Salmond G.P.C. 1994; Conserved virulence factor regulation and secretion systems in bacterial pathogens of plants and animals.. Eur J Plant Pathology 101:1–13
    [Google Scholar]
  24. Zhang Y., Broome-Smith J.K. 1990; Correct insertion of a simple eukaryotic plasma-membrane protein into the cytoplasmic membrane of Escherichia coli.. Gene 96:51–57
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-143-3-713
Loading
/content/journal/micro/10.1099/00221287-143-3-713
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error