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. 1986 Dec;168(3):1228–1233. doi: 10.1128/jb.168.3.1228-1233.1986

Molecular cloning and analysis of genes for production of K5, K7, K12, and K92 capsular polysaccharides in Escherichia coli.

I Roberts, R Mountford, N High, D Bitter-Suermann, K Jann, K Timmis, G Boulnois
PMCID: PMC213626  PMID: 3023300

Abstract

With a DNA fragment from within the region encoding the transport functions for K1 production as a hybridization probe in Southern blot experiments, homologous DNA sequences were detected in the DNA from Escherichia coli strains producing K5, K7, K92, and K100 capsular polysaccharides. No homology with the laboratory strain LE392 was detected. The same DNA probe was used to prescreen cosmid libraries in LE392 by colony hybridization, as a rapid method to isolate clones encoding the genes for K5, K7, K12, and K92 antigen production. Clones carrying sequences homologous to the probe that also produced capsular material were identified by using polyclonal and monoclonal antibodies raised against the K antigen in question and K antigen-specific phages. By restriction enzyme mapping of the appropriate cosmid clones it was possible to align the genes for the production of different K antigens in terms of common restriction endonuclease cleavage sites. A DNA fragment encoding the postulated transport functions for K7 antigen production could complement deletion mutations in the transport functions for K1 antigen production. Thus the transport to the cell surface of chemically distinct polysaccharides may be by a common process. Analysis in E. coli of the proteins produced by plasmids carrying the likely transport functions for K1, K5, and K7 antigen production revealed that each region coded for a similar polypeptide.

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Selected References

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

  1. Bhattacharjee A. K., Jennings H. J., Kenny C. P., Martin A., Smith I. C. Structural determination of the sialic acid polysaccharide antigens of Neisseria meningitidis serogroups B and C with carbon 13 nuclear magnetic resonance. J Biol Chem. 1975 Mar 10;250(5):1926–1932. [PubMed] [Google Scholar]
  2. Clewell D. B., Helinski D. R. Supercoiled circular DNA-protein complex in Escherichia coli: purification and induced conversion to an opern circular DNA form. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1159–1166. doi: 10.1073/pnas.62.4.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Collins J., Hohn B. Cosmids: a type of plasmid gene-cloning vector that is packageable in vitro in bacteriophage lambda heads. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4242–4246. doi: 10.1073/pnas.75.9.4242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Echarti C., Hirschel B., Boulnois G. J., Varley J. M., Waldvogel F., Timmis K. N. Cloning and analysis of the K1 capsule biosynthesis genes of Escherichia coli: lack of homology with Neisseria meningitidis group B DNA sequences. Infect Immun. 1983 Jul;41(1):54–60. doi: 10.1128/iai.41.1.54-60.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Egan W., Liu T. Y., Dorow D., Cohen J. S., Robbins J. D., Gotschlich E. C., Robbins J. B. Structural studies on the sialic acid polysaccharide antigen of Escherichia coli strain Bos-12. Biochemistry. 1977 Aug 9;16(16):3687–3692. doi: 10.1021/bi00635a028. [DOI] [PubMed] [Google Scholar]
  6. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  7. Finne J. Occurrence of unique polysialosyl carbohydrate units in glycoproteins of developing brain. J Biol Chem. 1982 Oct 25;257(20):11966–11970. [PubMed] [Google Scholar]
  8. Frosch M., Görgen I., Boulnois G. J., Timmis K. N., Bitter-Suermann D. NZB mouse system for production of monoclonal antibodies to weak bacterial antigens: isolation of an IgG antibody to the polysaccharide capsules of Escherichia coli K1 and group B meningococci. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1194–1198. doi: 10.1073/pnas.82.4.1194. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Glode M. P., Robbins J. B., Liu T. Y., Gotschlich E. C., Orskov I., Orskov F. Cross-antigenicity and immunogenicity between capsular polysaccharides of group C Neisseria meningitidis and of Escherichia coli K92. J Infect Dis. 1977 Jan;135(1):94–104. doi: 10.1093/infdis/135.1.94. [DOI] [PubMed] [Google Scholar]
  10. Gotschlich E. C., Fraser B. A., Nishimura O., Robbins J. B., Liu T. Y. Lipid on capsular polysaccharides of gram-negative bacteria. J Biol Chem. 1981 Sep 10;256(17):8915–8921. [PubMed] [Google Scholar]
  11. Grunstein M., Hogness D. S. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. doi: 10.1073/pnas.72.10.3961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hoffman S., Sorkin B. C., White P. C., Brackenbury R., Mailhammer R., Rutishauser U., Cunningham B. A., Edelman G. M. Chemical characterization of a neural cell adhesion molecule purified from embryonic brain membranes. J Biol Chem. 1982 Jul 10;257(13):7720–7729. [PubMed] [Google Scholar]
  13. Hohn B., Collins J. A small cosmid for efficient cloning of large DNA fragments. Gene. 1980 Nov;11(3-4):291–298. doi: 10.1016/0378-1119(80)90069-4. [DOI] [PubMed] [Google Scholar]
  14. Hohn B. In vitro packaging of lambda and cosmid DNA. Methods Enzymol. 1979;68:299–309. doi: 10.1016/0076-6879(79)68021-7. [DOI] [PubMed] [Google Scholar]
  15. Ichihara N., Ishimoto N., Ito E. Enzymatic incorporation of N-acetyl-D-mannosaminuronic acid and D-glucose into a polysaccharide of Escherichia coli 314 K7 H minus. FEBS Lett. 1974 Apr 1;40(2):309–311. doi: 10.1016/0014-5793(74)80251-6. [DOI] [PubMed] [Google Scholar]
  16. Jacobsson I., Lindahl U., Jensen J. W., Rodén L., Prihar H., Feingold D. S. Biosynthesis of heparin. Substrate specificity of heparosan N-sulfate D-glucuronosyl 5-epimerase. J Biol Chem. 1984 Jan 25;259(2):1056–1063. [PubMed] [Google Scholar]
  17. Kaijser B., Jodal U. Escherichia coli K5 antigen in relation to various infections and in healthy individuals. J Clin Microbiol. 1984 Feb;19(2):264–266. doi: 10.1128/jcm.19.2.264-266.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Liu T. Y., Gotschlich E. C., Dunne F. T., Jonssen E. K. Studies on the meningococcal polysaccharides. II. Composition and chemical properties of the group B and group C polysaccharide. J Biol Chem. 1971 Aug 10;246(15):4703–4712. [PubMed] [Google Scholar]
  19. MCGUIRE E. J., BINKLEY S. B. THE STRUCTURE AND CHEMISTRY OF COLOMINIC ACID. Biochemistry. 1964 Feb;3:247–251. doi: 10.1021/bi00890a017. [DOI] [PubMed] [Google Scholar]
  20. Navia J. L., Riesenfeld J., Vann W. F., Lindahl U., Rodén L. Assay of N-acetylheparosan deacetylase with a capsular polysaccharide from Escherichia coli K5 as substrate. Anal Biochem. 1983 Nov;135(1):134–140. doi: 10.1016/0003-2697(83)90741-8. [DOI] [PubMed] [Google Scholar]
  21. Orskov I., Orskov F., Jann B., Jann K. Serology, chemistry, and genetics of O and K antigens of Escherichia coli. Bacteriol Rev. 1977 Sep;41(3):667–710. doi: 10.1128/br.41.3.667-710.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Robbins J. B., Myerowitz L., Whisnant J. K., Argaman M., Schneerson R., Handzel Z. T., Gotschlich E. C. Enteric bacteria cross-reactive with Neisseria meningitidis groups A and C and Diplococcus pneumoniae types I and 3. Infect Immun. 1972 Nov;6(5):651–656. doi: 10.1128/iai.6.5.651-656.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. SAITO H., MIURA K. I. PREPARATION OF TRANSFORMING DEOXYRIBONUCLEIC ACID BY PHENOL TREATMENT. Biochim Biophys Acta. 1963 Aug 20;72:619–629. [PubMed] [Google Scholar]
  24. Schmidt M. A., Jann K. Structure of the 2-keto-3-deoxy-D-manno-octonic-acid-containing capsular polysaccharide (K12 antigen) of the urinary-tract-infective Escherichia coli O4:K12:H-. Eur J Biochem. 1983 Apr 5;131(3):509–517. doi: 10.1111/j.1432-1033.1983.tb07291.x. [DOI] [PubMed] [Google Scholar]
  25. Silver R. P., Finn C. W., Vann W. F., Aaronson W., Schneerson R., Kretschmer P. J., Garon C. F. Molecular cloning of the K1 capsular polysaccharide genes of E. coli. Nature. 1981 Feb 19;289(5799):696–698. doi: 10.1038/289696b0. [DOI] [PubMed] [Google Scholar]
  26. Silver R. P., Vann W. F., Aaronson W. Genetic and molecular analyses of Escherichia coli K1 antigen genes. J Bacteriol. 1984 Feb;157(2):568–575. doi: 10.1128/jb.157.2.568-575.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  28. Timmis K. N., Boulnois G. J., Bitter-Suermann D., Cabello F. C. Surface components of Escherichia coli that mediate resistance to the bactericidal activities of serum and phagocytes. Curr Top Microbiol Immunol. 1985;118:197–218. doi: 10.1007/978-3-642-70586-1_11. [DOI] [PubMed] [Google Scholar]
  29. Troy F. A., 2nd The chemistry and biosynthesis of selected bacterial capsular polymers. Annu Rev Microbiol. 1979;33:519–560. doi: 10.1146/annurev.mi.33.100179.002511. [DOI] [PubMed] [Google Scholar]
  30. Tsui F. P., Boykins R. A., Egan W. Structural and immunological studies of the Escherichia coli K7 (K56) capsular polysaccharide. Carbohydr Res. 1982 Apr 16;102:263–271. doi: 10.1016/s0008-6215(00)88068-4. [DOI] [PubMed] [Google Scholar]
  31. Vann W. F., Schmidt M. A., Jann B., Jann K. The structure of the capsular polysaccharide (K5 antigen) of urinary-tract-infective Escherichia coli 010:K5:H4. A polymer similar to desulfo-heparin. Eur J Biochem. 1981 May 15;116(2):359–364. doi: 10.1111/j.1432-1033.1981.tb05343.x. [DOI] [PubMed] [Google Scholar]
  32. Wilkins B. M., Boulnois G. J., Lanka E. A plasmid DNA primase active in discontinuous bacterial DNA replication. Nature. 1981 Mar 19;290(5803):217–221. doi: 10.1038/290217a0. [DOI] [PubMed] [Google Scholar]

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