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. 1985 Sep;50(3):562–567. doi: 10.1128/aem.50.3.562-567.1985

Partial purification and characterization of a polymannuronic acid depolymerase produced by a mucoid strain of Pseudomonas aeruginosa isolated from a patient with cystic fibrosis.

W M Dunne Jr, F L Buckmire
PMCID: PMC238669  PMID: 3935048

Abstract

An exopolysaccharide depolymerase was isolated from a mucoid strain of Pseudomonas aeruginosa of cystic fibrosis origin. Purified preparations of the depolymerase showed maximum activity against the unacetylated polymannuronic acid exopolysaccharide from the same strain and little activity against commercially prepared alginic acid. The evidence suggests that the enzyme is either periplasmic in location or associated with the outer cell membrane and is released extracellularly, in the absence of cell lysis, after a reduction of the culture magnesium (Mg2+) concentration below 3.0 mM. The depolymerase is also released after the addition of sublethal concentrations of EDTA to cultures containing 3.0 mM Mg2+. A survey of additional mucoid P. aeruginosa isolates recovered from patients with cystic fibrosis showed that nearly 60% demonstrated similar depolymerase activity while none of the nonmucoid revertants of the parent strains produced detectable depolymerase activity.

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Selected References

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  1. Boyd J., Turvey J. R. Isolation of poly-alpha-L-guluronate lyase from Klebsiella aerogenes. Carbohydr Res. 1977 Aug;57:163–171. doi: 10.1016/s0008-6215(00)81928-x. [DOI] [PubMed] [Google Scholar]
  2. Bucke C. Polyacrylamide gel electrophoresis of alginic acid. J Chromatogr. 1974 Feb 13;89(1):99–102. doi: 10.1016/s0021-9673(01)84167-4. [DOI] [PubMed] [Google Scholar]
  3. CETIN E. T., TOERECI K., ANG O. ENCAPSULATED PSEUDOMONAS AERUGINOSA (PSEUDOMONAS AERUGINOSA MUCOSUS) STRAINS. J Bacteriol. 1965 May;89:1432–1433. doi: 10.1128/jb.89.5.1432-1433.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carlson D. M., Matthews L. W. Polyuronic acids produced by Pseudomonas aeruginosa. Biochemistry. 1966 Sep;5(9):2817–2822. doi: 10.1021/bi00873a006. [DOI] [PubMed] [Google Scholar]
  5. Cheng K. J., Ingram J. M., Costerton J. W. Alkaline phosphatase localization and spheroplast formation of Pseudomonas aeruginosa. Can J Microbiol. 1970 Dec;16(12):1319–1324. doi: 10.1139/m70-218. [DOI] [PubMed] [Google Scholar]
  6. Cheng K. J., Ingram J. M., Costerton J. W. Release of alkaline phosphatase from cells of Pseudomonas aeruginosa by manipulation of cation concentration and of pH. J Bacteriol. 1970 Nov;104(2):748–753. doi: 10.1128/jb.104.2.748-753.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davidson I. W., Sutherland I. W., Lawson C. J. Purification and properties of an alginate lyase from a marine bacterium. Biochem J. 1976 Dec 1;159(3):707–713. doi: 10.1042/bj1590707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Doggett R. G. Incidence of mucoid Pseudomonas aeruginosa from clinical sources. Appl Microbiol. 1969 Nov;18(5):936–937. doi: 10.1128/am.18.5.936-937.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Doubet R. S., Quatrano R. S. Properties of alginate lyases from marine bacteria. Appl Environ Microbiol. 1984 Apr;47(4):699–703. doi: 10.1128/aem.47.4.699-703.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Elston H. R., Hoffman K. C. Increasing incidence of encapsulated Pseudomonas aeruginosa strains. Am J Clin Pathol. 1967 Nov;48(5):519–523. doi: 10.1093/ajcp/48.5_ts.519. [DOI] [PubMed] [Google Scholar]
  11. Evans L. R., Linker A. Production and characterization of the slime polysaccharide of Pseudomonas aeruginosa. J Bacteriol. 1973 Nov;116(2):915–924. doi: 10.1128/jb.116.2.915-924.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Govan J. R., Fyfe J. A. Mucoid Pseudomonas aeruginosa and cystic fibrosis: resistance of the mucoid from to carbenicillin, flucloxacillin and tobramycin and the isolation of mucoid variants in vitro. J Antimicrob Chemother. 1978 May;4(3):233–240. doi: 10.1093/jac/4.3.233. [DOI] [PubMed] [Google Scholar]
  13. Hancock R. E., Raffle V. J., Nicas T. I. Involvement of the outer membrane in gentamicin and streptomycin uptake and killing in Pseudomonas aeruginosa. Antimicrob Agents Chemother. 1981 May;19(5):777–785. doi: 10.1128/aac.19.5.777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hansen J. B., Doubet R. S., Ram J. Alginase enzyme production by Bacillus circulans. Appl Environ Microbiol. 1984 Apr;47(4):704–709. doi: 10.1128/aem.47.4.704-709.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Haug A., Larsen B. Biosynthesis of alginate. II. Polymannuronic acid C-5-epimerase from Azotobacter vinelandii (Lipman). Carbohydr Res. 1971 Apr;17(2):297–308. doi: 10.1016/s0008-6215(00)82537-9. [DOI] [PubMed] [Google Scholar]
  16. LINKER A., JONES R. S. A POLYSACCHARIDE RESEMBLING ALGINIC ACID FROM A PSEUDOMONAS MICRO-ORGANISM. Nature. 1964 Oct 10;204:187–188. doi: 10.1038/204187a0. [DOI] [PubMed] [Google Scholar]
  17. Lam J., Chan R., Lam K., Costerton J. W. Production of mucoid microcolonies by Pseudomonas aeruginosa within infected lungs in cystic fibrosis. Infect Immun. 1980 May;28(2):546–556. doi: 10.1128/iai.28.2.546-556.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Laraya-Cuasay L. R., Cundy K. R., Huang N. N. Pseudomonas carrier rates of patients with cystic fibrosis and of members of their families. J Pediatr. 1976 Jul;89(1):23–26. doi: 10.1016/s0022-3476(76)80920-1. [DOI] [PubMed] [Google Scholar]
  19. Larsen B., Haug A. Biosynthesis of alginate. 1. Composition and structure of alginate produced by Azotobacter vinelandii (Lipman). Carbohydr Res. 1971 Apr;17(2):287–296. doi: 10.1016/s0008-6215(00)82536-7. [DOI] [PubMed] [Google Scholar]
  20. Larsen B., Haug A. Biosynthesis of alginate. 3. Tritium incorporation with polymannuronic acid 5-epimerase from Azotobacter vinelandii. Carbohydr Res. 1971 Dec;20(2):225–232. doi: 10.1016/s0008-6215(00)81375-0. [DOI] [PubMed] [Google Scholar]
  21. Linker A., Jones R. S. A new polysaccharide resembling alginic acid isolated from pseudomonads. J Biol Chem. 1966 Aug 25;241(16):3845–3851. [PubMed] [Google Scholar]
  22. Min K. H., Sasaki S., Kashiwabara Y., Nisizawa K. Substrate specificity of endo-polyguluronide lyases from Pseudomonas sp. on the basis of their kinetic properties. J Biochem. 1977 Mar;81(3):547–553. doi: 10.1093/oxfordjournals.jbchem.a131489. [DOI] [PubMed] [Google Scholar]
  23. PREISS J., ASHWELL G. Alginic acid metabolism in bacteria. II. The enzymatic reduction of 4-deoxy-L-erythro-5-hexoseulose uronic acid to 2-keto-3-deoxy-D-gluconic acid. J Biol Chem. 1962 Feb;237:317–321. [PubMed] [Google Scholar]
  24. Schnaitman C. A. Comparison of rat liver mitochondrial and microsomal membrane proteins. Proc Natl Acad Sci U S A. 1969 Jun;63(2):412–419. doi: 10.1073/pnas.63.2.412. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Schwarzmann S., Boring J. R. Antiphagocytic Effect of Slime from a Mucoid Strain of Pseudomonas aeruginosa. Infect Immun. 1971 Jun;3(6):762–767. doi: 10.1128/iai.3.6.762-767.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. WEISSBACH A., HURWITZ J. The formation of 2-keto-3-deoxyheptonic acid in extracts of Escherichia coli B. I. Identification. J Biol Chem. 1959 Apr;234(4):705–709. [PubMed] [Google Scholar]
  27. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  28. di Sant'Agnese P. A., Davis P. B. Research in cystic fibrosis (third of three parts). N Engl J Med. 1976 Sep 9;295(11):597–602. doi: 10.1056/NEJM197609092951105. [DOI] [PubMed] [Google Scholar]

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