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. 1988 Sep;170(9):4008–4014. doi: 10.1128/jb.170.9.4008-4014.1988

Accumulation of a lipid-linked intermediate involved in enterobacterial common antigen synthesis in Salmonella typhimurium mutants lacking dTDP-glucose pyrophosphorylase.

P D Rick 1, S Wolski 1, K Barr 1, S Ward 1, L Ramsay-Sharer 1
PMCID: PMC211403  PMID: 2842298

Abstract

The heteropolysaccharide chains of enterobacterial common antigen (ECA) are composed of linear trisaccharide repeat units having the structure----3)-alpha-Fuc4NAc-(1----4)-beta-D-ManNAcA-(1---- 4)-alpha-D-GlcNAc- (1----. Mutants of Salmonella typhimurium lacking the structural gene for dTDP-glucose pyrophosphorylase (rfbA) are severely impaired in their ability to synthesize dTDP-glucose, which is a precursor of dTDP-4-acetamido-4,6-dideoxy-D-galactose (Fuc4NAc), the donor of Fuc4NAc residues for ECA synthesis. These mutants synthesize only trace amounts of ECA, and they are hypersensitive to sodium dodecyl sulfate (SDS). Incubation of delta rfbA mutants with [3H]N-acetylglucosamine ([3H]GlcNAc) resulted in the accumulation of radioactivity in N-acetyl-D-mannosaminuronic acid (ManNAcA)-GlcNAc-pyrophosphorylundecaprenol (lipid II), the putative acceptor of Fuc4NAc residues in ECA synthesis. Lipid II did not accumulate in either wild-type cells or in rff mutants unable to synthesize ManNAcA. Both the accumulation of lipid II and the synthesis of trace amounts of ECA were abolished when delta rfbA mutants were grown in the presence of the antibiotic tunicamycin. Tunicamycin also prevented the SDS-mediated lysis of the mutants. SDS-resistant derivatives of delta rfbA mutants were isolated that were no longer able to synthesize trace amounts of ECA. Characterization of these derivatives revealed that they were defective in various steps of ECA synthesis leading to the synthesis of lipid II. The data support the conclusion that accumulation of lipid II is responsible in some way for the hypersensitivity of delta rfbA mutants to SDS.

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Selected References

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  1. Barr K., Rick P. D. Biosynthesis of enterobacterial common antigen in Escherichia coli. In vitro synthesis of lipid-linked intermediates. J Biol Chem. 1987 May 25;262(15):7142–7150. [PubMed] [Google Scholar]
  2. Basu S., Kuhn H. M., Neszmelyi A., Himmelspach K., Mayer H. Chemical characterization of enterobacterial common antigen isolated from Plesiomonas shigelloides ATCC 14029. Eur J Biochem. 1987 Jan 2;162(1):75–81. doi: 10.1111/j.1432-1033.1987.tb10544.x. [DOI] [PubMed] [Google Scholar]
  3. Ichihara N., Ishimoto N., Ito E. Enzymatic formation of uridine diphosphate N-acetyl-D-mannosaminuronic acid. FEBS Lett. 1974 Feb 1;39(1):46–48. doi: 10.1016/0014-5793(74)80013-x. [DOI] [PubMed] [Google Scholar]
  4. Kuhn H. M., Neter E., Mayer H. Modification of the lipid moiety of the enterobacterial common antigen by the "Pseudomonas factor". Infect Immun. 1983 May;40(2):696–700. doi: 10.1128/iai.40.2.696-700.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  6. Lew H. C., Mäkelä P. H., Kuhn H. M., Mayer H., Nikaido H. Biosynthesis of enterobacterial common antigen requires dTDPglucose pyrophosphorylase determined by a Salmonella typhimurium rfb gene and a Salmonella montevideo rfe gene. J Bacteriol. 1986 Nov;168(2):715–721. doi: 10.1128/jb.168.2.715-721.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Lew H. C., Nikaido H., Mäkelä P. H. Biosynthesis of uridine diphosphate N-acetylmannosaminuronic acid in rff mutants of Salmonella tryphimurium. J Bacteriol. 1978 Oct;136(1):227–233. doi: 10.1128/jb.136.1.227-233.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Mayer H., Schmidt G. Chemistry and biology of the enterobacterial common antigen (ECA). Curr Top Microbiol Immunol. 1979;85:99–153. doi: 10.1007/978-3-642-67322-1_3. [DOI] [PubMed] [Google Scholar]
  9. Meier U., Mayer H. Genetic location of genes encoding enterobacterial common antigen. J Bacteriol. 1985 Aug;163(2):756–762. doi: 10.1128/jb.163.2.756-762.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Mäkelä P. H., Jahkola M., Lüderitz O. A new gene cluster rfe concerned with the biosynthesis of Salmonella lipopolysaccharide. J Gen Microbiol. 1970 Jan;60(1):91–106. doi: 10.1099/00221287-60-1-91. [DOI] [PubMed] [Google Scholar]
  11. Mäkelä P. H., Mayer H. Enterobacterial common antigen. Bacteriol Rev. 1976 Sep;40(3):591–632. doi: 10.1128/br.40.3.591-632.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mäkelä P. H. Participation of lipopolysaccharide genes in the determination of the entobacterial common antigen: analysis in Salmonella groups B and C1. J Bacteriol. 1974 Sep;119(3):765–770. doi: 10.1128/jb.119.3.765-770.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mäkelä P. H., Schmidt G., Mayer H., Nikaido H., Whang H. Y., Neter E. Enterobacterial common antigen in rfb deletion mutants of Salmonella typhimurium. J Bacteriol. 1976 Sep;127(3):1141–1149. doi: 10.1128/jb.127.3.1141-1149.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. Peters H., Jürs M., Jann B., Jann K., Timmis K. N., Bitter-Suermann D. Monoclonal antibodies to enterobacterial common antigen and to Escherichia coli lipopolysaccharide outer core: demonstration of an antigenic determinant shared by enterobacterial common antigen and E. coli K5 capsular polysaccharide. Infect Immun. 1985 Nov;50(2):459–466. doi: 10.1128/iai.50.2.459-466.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. ROTHFIELD L., OSBORN M. J., HORECKER B. L. BIOSYNTHESIS OF BACTERIAL LIPOPOLYSACCHARIDE. II. INCORPORATION OF GLUCOSE AND GALACTOSE CATALYZED BY PARTICULATE AND SOLUBLE ENZYMES IN SALMONELLA. J Biol Chem. 1964 Sep;239:2788–2795. [PubMed] [Google Scholar]
  17. Rick P. D., Mayer H., Neumeyer B. A., Wolski S., Bitter-Suermann D. Biosynthesis of enterobacterial common antigen. J Bacteriol. 1985 May;162(2):494–503. doi: 10.1128/jb.162.2.494-503.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Rick P. D., Osborn M. J. Lipid A mutants of Salmonella typhimurium. Characterization of a conditional lethal mutant in 3-deoxy-D-mannooctulosonate-8-phosphate synthetase. J Biol Chem. 1977 Jul 25;252(14):4895–4903. [PubMed] [Google Scholar]
  19. Rick P. D., Young D. A. Isolation and characterization of a temperature-sensitive lethal mutant of Salmonella typhimurium that is conditionally defective in 3-deoxy-D-manno-octulosonate-8-phosphate synthesis. J Bacteriol. 1982 May;150(2):447–455. doi: 10.1128/jb.150.2.447-455.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sanderson K. E., Ross H., Ziegler L., Mäkelä P. H. F + , Hfr, and F' strains of Salmonella typhimurium and Salmonella abony. Bacteriol Rev. 1972 Dec;36(4):608–637. doi: 10.1128/br.36.4.608-637.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schmidt G., Mayer H., Mäkelä P. H. Presence of rfe genes in Escherichia coli: their participation in biosynthesis of O antigen and enterobacterial common antigen. J Bacteriol. 1976 Aug;127(2):755–762. doi: 10.1128/jb.127.2.755-762.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Yuasa R., Levinthal M., Nikaido H. Biosynthesis of cell wall lipopolysaccharide in mutants of Salmonella. V. A mutant of Salmonella typhimurium defective in the synthesis of cytidine diphosphoabequose. J Bacteriol. 1969 Oct;100(1):433–444. doi: 10.1128/jb.100.1.433-444.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]

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