1887

Abstract

SUMMARY: Biosynthesis of the Escherichis coli K5 polysaccharide requires the Kf iA, Kf iB, KfiC and KfiD proteins. The subsequent transport of the polysaccharide onto the cell surface requires the KpsC, KpsD, KpsE, KpsM, KpsS and KpsT proteins, which are conserved between different group II capsular polysaccharides. The KfiA and KfiC, together with the KpsC, KpsS and KpsT proteins, were purified and polyclonal antisera to each protein generated. These antisera, together with one previously generated (by others) against the purified KfiD protein, were used in Western blot analysis to locate the corresponding proteins within the cell. Analysis of membrane fractions revealed that KfiA (involved in initiation of polysaccharide synthesis), Kf iC (K5 glycosyl transferase) and the Kf iD protein (UDP-glucose dehydrogenase) were associated with the inner membrane. The KpsC, KpsS and KpsT proteins involved in polysaccharide transport were associated with the inner membrane and this membrane association occurred in the absence of any other capsule-related proteins. The effect of mutations in individual kps genes on the localization of each protein was determined. Mutations in the kpC# kpsM, kpsS and kpsT genes resulted in a loss of membrane targeting for KfiA and KfiC, suggesting some form of hetero-oligomeric membrane-bound biosynthetic complex. Osmotic shock caused the release of KfiA, KfiC, KpsC and KpsS from the inner membrane into the periplasm, suggesting that the polysaccharide biosynthetic complex may be associated with sites of adhesion between the inner and outer membrane.

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1998-10-01
2021-05-18
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References

  1. Bayer M.E. 1991; Zones of membrane adhesion in the cryofixed envelope of Escherichia coli.. J Struct Biol 107:268–280
    [Google Scholar]
  2. Bayer M.E., Thurow H. 1977; Polysaccharide capsule of Escherichia coli: microscopy study of its size, structure, and sites of synthesis.. J Bacteriol 130:911–936
    [Google Scholar]
  3. Bayer M.E., Bayer M.H., Lunn C.A., Pigiet V. 1987; Association of thioredoxin with the inner membrane and adhesion sites in Escherichia coli.. J Bacteriol 169:2659–2666
    [Google Scholar]
  4. Beacham I.R. 1979; Periplasmic enzymes in Gram-negative bacteria.. Int J Biochem 10:877–883
    [Google Scholar]
  5. Bliss J.M., Silver R. 1997; Evidence that KpsT, the ATP- binding component of an ATP-binding casette transporter, is exposed to the periplasm and associates with polymer during translocation of the polysialic acid capsule of Escherichia coli Kl.. J Bacteriol 179:1400–1403
    [Google Scholar]
  6. Boulnois G.J., Roberts I.S. 1990; Genetics of capsular polysaccharide production in bacteria.. Curr Top Microbiol Immunol 150:1–18
    [Google Scholar]
  7. Bronner D., Sieberth V., Pazzani C., Smith A.S., Boulnois G.J., Roberts I.S., Jann B., Jann K. 1993a; Synthesis of the K5 (group 2) capsular polysaccharide in transport-deficient recombinant Escherichia coli.. FEMS Microbiol Lett 113:279–284
    [Google Scholar]
  8. Bronner D., Sieberth V., Pazzani C., Roberts I.S., Boulnois G.J., Jann B., Jann K. 1993b; Expression of the capsular K5 polysaccharide of Escherichia coli: biochemical and electron microscopic analyses of mutants with defects in region 1 of the K5 gene cluster.. J Bacteriol 175:5984–5992
    [Google Scholar]
  9. Chang J.Y., Haller B., Fuchs J.A. 1985; Thioredoxin is the bacterial protein encoded by ftp that is required for filamentous bacteriophage assembly.. J Bacteriol 161:799–802
    [Google Scholar]
  10. Cieslewicz M.J., Vimr E.R. 1997; Reduced sialic acid capsule expression in Escherichia coli Kl mutants with defects in kpsF.. Mol Microbiol 26:237–249
    [Google Scholar]
  11. Earhart C.F., Hantash F. 1995; Osmotic shock-sensitive proteins of the Escherichia coli enterobactin synthase complex.. J Cell Biochem S21A:244
    [Google Scholar]
  12. El Yaagoubi A., Kohiyama M., Richarme G. 1994; Localisation of DnaK (chaperone 70) from Escherichia coli in an osmotic sensitive compartment of the cytoplasm.. J Bacteriol 176:7074–7078
    [Google Scholar]
  13. Finke A., Bronner B., Nikolaev A.V., Jann B., Jann K. 1991; Biosynthesis of the Escherichia coli K5 polysaccharide, a representative of group II capsular polysaccharides: polymerisation in vitro and characterisation of the product.. J Bacteriol 173:4088–4094
    [Google Scholar]
  14. Frosch M., Müller A.M. 1993; Phospholipid substitution of capsular polysaccharides and mechanism of capsule formation in Neisseria meningitidis.. Mol Microbiol 8:483–493
    [Google Scholar]
  15. Gransden W., Eykyn S., Phillips I., Rowe B. 1990; Bacteremia due to Escherichia coli: a study of 861 episodes.. Rev Infect Dis 12:1008–1018
    [Google Scholar]
  16. Griffiths G., Cook N.J., Gottfridson E., Lind T., Lidholt K., Roberts I.S. 1998; Characterization of the glycosyltransferase enzyme from the Escherichia coli K5 capsule gene cluster and identification and characterization of the glucuronyl active site.. J Biol Chem 273:11752–11757
    [Google Scholar]
  17. Guihard G., Boulanger P., Bénédetti H., Loubés R., Bernard M., Letellier L. 1994; Colicin A and Tol proteins involved in its translocation are preferentially located in the contact sites between the inner and outer membranes of Escherichia coli cells.. J Biol Chem 269:5874–5880
    [Google Scholar]
  18. Hantash F.M., Ammerlaan M., Earhart C.F. 1997; Entero-bactin synthase polypeptides of Escherichia coli are present in an osmotic-shock-sensitive cytoplasmic locality.. Microbiology 143:147–156
    [Google Scholar]
  19. Harlow E., Lane D. 1988 Antibodies, a Laboratory Manual. Chapter 15 pp. 613–633 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  20. Hayashi M., Miyoshi T., Takashina S., Unemoto T. 1989; Purification of NADH-ferricyanide dehydrogenase and NADH- quinone reductase from Escherichia coli membranes and their roles in the respiratory chain.. Biochim Biophys Acta 977:62–69
    [Google Scholar]
  21. von Heijne G. 1994; Membrane proteins: from sequence to structure.. Annu Rev Biophys Biomol Struct 23:167–192
    [Google Scholar]
  22. High N.J., Hales B.A., Jann K., Boulnois G.J. 1988; A block of urovirulence genes encoding multiple fimbriae and haemolysin in Escherichia coli 04:K12:H-.. Infect Immun 56:513–517
    [Google Scholar]
  23. Hu G. 1993; DNA polymerase-catalyzed addition of non- templated extra nucleotides to the 3’ end of a DNA fragment.. DNA Cell Biol 12:763–770
    [Google Scholar]
  24. Jann B., Jann K. 1990; Structure and biosynthesis of the capsular antigens of Escherichia coli.. Curr Top Microbiol Immunol 150:19–42
    [Google Scholar]
  25. Jann K., Jann B. 1992; Capsules of Escherichia coli, expression and biological significance.. Can J Microbiol 38:705–710
    [Google Scholar]
  26. Lunn C.A., Pigiet V. 1986; Localisation of thioredoxin in Escherichia coli. . In Thioredoxin and Glutaredoxin Systems: Structure and Function pp. 165–176 Holmgren A. others Edited by New York: Raven Press;
    [Google Scholar]
  27. de Maagd R.A., Lugtenberg B. 1986; Fractionation of Rhizobium leguminosarum cells into outer membrane, cytoplasmic membrane, periplasmic and cytoplasmic components.. J Bacteriol 167:1083–1085
    [Google Scholar]
  28. Pavelka M.S., Hayes S.F., Silver R.P. 1994; Characterisation of KpsT, the ATP-binding component of the ABC-transporter involved in the export of capsular polysialic acid in Escherichia coli Kl.. J Biol Chem 269:20149–20158
    [Google Scholar]
  29. Pazzani C. 1992 Molecular and genetic analysis of the Escherichia coli K5 capsule gene cluster. PhD thesis University of Leiceste.;
    [Google Scholar]
  30. Pazzani C., Rosenow C., Boulnois G.J., Bronner D., Jann K., Roberts I.S. 1993; Molecular analysis of region 1 of the Escherichia coli capsule gene cluster: a region encoding proteins involved in cell surface expression of capsular polysaccharide.. J Bacteriol 175:5978–5983
    [Google Scholar]
  31. Pearce R., Roberts I.S. 1995; Cloning and analysis of gene clusters for production of the Escherichia coli K10 and K54 antigens: identification of a new group of serA-linked capsule gene clusters.. J Bacteriol 177:3992–3997
    [Google Scholar]
  32. Petit C., Rigg G.P., Pazzani C., smith A., Sieberth V., Stevens M., Boulnois G.J., Jann K., Roberts I.S. 1995; Region 2 of the Escherichia coli K5 capsule gene cluster encoding proteins for the biosynthesis of the K5 polysaccharide.. Mol Microbiol 17:611–620
    [Google Scholar]
  33. Pigeon R.P., Silver R.P. 1994; Topological and mutational analysis of KpsM, the hydrophobic component of the ABC-transporter involved in the export of polysialic acid in Escherichia coli Kl.. Mol Microbiol 14:871–881
    [Google Scholar]
  34. Rheus B.R., Williams M.N.V., Kim J.S., Carlson R.W., Cõté F. 1995; Suppression of the Fix“ phenotype of Rhizobium meliloti exoB mutants by LpsZ is correlated to a modified expression of the K polysaccharide.. J Bacteriol 177:4289–4296
    [Google Scholar]
  35. Roberts I.S. 1995; Bacterial capsules in sickness and in health.. Microbiology 141:2023–2031
    [Google Scholar]
  36. Roberts I.S. 1996; Biochemistry and genetics of capsular polysaccharide production in bacteria.. Annu Rev Microbiol 50:285–315
    [Google Scholar]
  37. Roberts I.S., Mountford R., High N., Bitter-Suerman D., Jann K., Timmis K.N., Boulnois G.J. 1986; Molecular cloning and analysis of genes for production of K5, K7, K12, and K92 capsular polysaccharides in Escherichia coli.. J Bacteriol 170:1228–1233
    [Google Scholar]
  38. Roberts I.S., Mountford R., Hodge R., Jann K., Boulnois G.J. 1988; Common organisation of gene clusters of different capsular polysaccharides (K antigens) from Escherichia coli.. J Bacteriol 170:1305–1310
    [Google Scholar]
  39. Roberts M., Roberts I.S, Korhonen T., Jann K., Bitter-Suerman D., Boulnois G.J., Williams P.H. 1988; DNA probes for K antigen (capsule) typing of Escherichia coli.. J Clin Microbiol 26:385–387
    [Google Scholar]
  40. Rosenow C., Esumeh F., Roberts I.S., Jann K. 1995a; Characterisation and localisation of the KpsE protein of Escherichia coli K5, which is involved in polysaccharide export.. J Bacteriol 117:1137–1143
    [Google Scholar]
  41. Rosenow C., Roberts I.S., Jann K. 1995b; Isolation from recombinant Escherichia coli and characterisation of CMP-Kdo synthetase, involved in the expression of the capsular K5 polysaccharide (K-CKS).. FEMS Microbiol Lett 125:159–164
    [Google Scholar]
  42. Saiki R.K., Gyllensten U.B., Erlich H. 1988 The Polymerase Chain Reaction in Genome Analysis. Davies K.E. Edited by Oxford: IRL Press;
    [Google Scholar]
  43. 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]
  44. Schmidt A.M., Jann K. 1982; Phospholipid substitution of capsular (K) polysaccharide antigens from Escherichia coli causing intestinal infections.. FEMS Microbiol Lett 14:69–74
    [Google Scholar]
  45. Sieberth V., Rigg G.P., Roberts I.S., Jann K. 1995; Expression and characterisation of UDPGlc dehydrogenase (KfiD), which is encoded in the type-specific region 2 of the Escherichia coli K5 capsule genes.. J Bacteriol 177:4562–4565
    [Google Scholar]
  46. Simpson D.A., Hammarton T.C., Roberts I.S. 1996; Trans-criptional organisation and regulation of expression of the Escherichia coli K5 capsule gene cluster.. J Bacteriol 178:6466–6474
    [Google Scholar]
  47. Smith A.N., Boulnois G.J., Roberts I.S. 1990; Molecular analysis of the Escherichia coli K5 kps locus: identification and characterisation of an inner-membrane capsular polysaccharide transport system.. Mol Microbiol 4:1863–1869
    [Google Scholar]
  48. Stevens M., Hanfling P., Jann B., Jann K., Roberts I.S. 1994; Regulation of Escherichia coli K5 capsular polysaccharide expression : evidence for involvement of RfaH in the expression of group II capsules.. FEMS Microbiol Lett 124:93–98
    [Google Scholar]
  49. Stevens M.P., Clarke B.R., Roberts I.S. 1997; Regulation of the Escherichia coli K5 capsule gene cluster by transcription antitermination.. Mol Microbiol 24:1001–1012
    [Google Scholar]
  50. Whitfield C., Vimr E.R., Costerton J.W., Troy F.A. 1985; Membrane proteins correlated with expression of polysialic acid capsule in Escherichia coli Kl.. J Bacteriol 161:743–749
    [Google Scholar]
  51. Wooldridge K.G., Morissey J.A., Williams P.H. 1992; Transport of ferric-aerobactin into the cytoplasm of Escherichia coli K12: role of envelope-associated proteins and effect of endogenous siderophores.. J Gen Microbiol 138:597–603
    [Google Scholar]
  52. Wunder D., Aaronson W., Hayes S., Bliss J., Silver R.P. 1994; Nucleotide sequence and mutational analysis of the gene encoding KpsD, a periplasmic protein involved in the transport of polysialic acid in Escherichia coli Kl.. J Bacteriol 176:4025–4033
    [Google Scholar]
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