Advertisement

Current Microbiology

, Volume 20, Issue 6, pp 359–363 | Cite as

Isolation of an extracellular polysaccharide (EPS) depolymerase produced byBradyrhizobium japonicum

  • Michael F. Dunn
  • Arthur L. Karr
Article

Abstract

Bradyrhizobium japonicum is capable of producing an acidic, high molecular weight, extracellular polysaccharide (EPS). An enzyme exhibiting EPS depolymerase activity was detected in cell lysates ofB. japonicum strain 2143. The depolymerase was active against the EPS produced by strain 2143 and the closely related EPS produced by strain 311b 110. Depolymerase activity was characterized by its ability to decrease the viscosity of EPS solutions, to decrease the molecular weight of EPS, and to catalyze the release of reducing groups from EPS. The depolymerase exhibited a sharp activity optimum at pH 6 and had a molecular weight of approximately 45 kD as determined by gel permeation chromatography. Analysis of depolymerase-treated EPS indicates that the enzyme acts as an endo-depolymerase, producing a relatively narrow size range of high molecular weight products.

Keywords

Enzyme Viscosity Molecular Weight Polysaccharide High Molecular Weight 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Literature Cited

  1. 1.
    Borthakur D, Downie JA, Johnson AWB, Lamb JW (1985)psi, a plasmid-linked gene that inhibits exopolysaccharide production and which is required for symbiotic nitrogen fixation. Mol Gen Genet 200:278–282Google Scholar
  2. 2.
    Borthakur D, Barber CE, Lamb JW, Daniels MJ, Downie JA, Johnson AWB (1986) A mutation that blocks exopolysaccharide synthesis prevents nodulation of peas byRhizobium leguminosarum but not of beans byR. phaseoli and is corrected by cloned DNA fromRhizobium or the phytopathogenXanthomonas. Mol Gen Genet 203:320–323Google Scholar
  3. 3.
    Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254PubMedGoogle Scholar
  4. 4.
    Cangleosi GA, Hung L, Puvanesarajah V, Stacey G, Ozga DA, Leigh JA, Nester EW (1987) Common loci forAgrobacterium tumefaciens andRhizobium meliloti exopolysaccharide synthesis and their roles in plant interactions. J Bacteriol 169:2086–2091PubMedGoogle Scholar
  5. 5.
    Chen H, Batley M, Redmond J, Rolfe BG (1985) Alteration of the effective nodulation properties of a fast-growing broad host rangeRhizobium due to changes in exopolysaccharide synthesis. J Plant Physiol 120:331–349Google Scholar
  6. 6.
    Davidson IW, Sutherland IW, Lawson CJ (1976) Purification and properties of an alginate lyase from a marine bacterium. Biochem J 159:707–713PubMedGoogle Scholar
  7. 7.
    Davis KR, Lyon GD, Darvill AG, Albersheim P (1984) Host-pathogen interactions. XXV. Endopolygalacturonic acid lyase fromErwinia carotovora elicits phytoalexin accumulation by releasing plant cell wall fragments. Plant Physiol 74:52–60Google Scholar
  8. 8.
    Dische, Z (1962) Color reactions of carbohydrates. In Whistler RL, Wolfram ML (eds) Methods in Carbohydrate Chemistry, Vol. 1. New York: Academic Press, pp 478–481Google Scholar
  9. 9.
    Djordjevic SP, Chen H, Batley M, Redmond JW, Rolfe BG (1987) Nitrogen fixation ability of exopolysaccharide synthesis mutants ofRhizobium sp. strain NGR234 andRhizobium trifolii is restored by the addition of homologous exopolysac-charides. J Bacteriol 169:53–60PubMedGoogle Scholar
  10. 10.
    Dudley ME, Long SR (1989) A non-nodulating alfalfa mutant displays neither root hair curling nor early cell division in response toRhizobium meliloti. Plant Cell 1:65–72PubMedGoogle Scholar
  11. 11.
    Dudman WF (1978) Structural studies of the extracellular polysaccharides ofRhizobium japonicum strains 71A, CC708 and CB 1795. Carbohydr Res 66:9–23Google Scholar
  12. 12.
    Dunne WM, Buckmire FLA (1985) Partial purification and characterization of a polymannuronic acid depolymerase produced by a mucoid strain ofPseudomonas aeruginosa isolated from a patient with cystic fibrosis. Appl Environ Microbiol 50:562–567PubMedGoogle Scholar
  13. 13.
    Dygart S, Li LH, Florida D, Thoma JA (1965) Determination of reducing sugar with improved precision. Anal Biochem 13:367–374PubMedGoogle Scholar
  14. 14.
    Eberhard S, Doubrava N, Marfa V, Mohnen D, Southwick A, Darvill A, Albersheim P (1989) Pectic cell wall fragments regulate tobacco thin-cell-layer explant morphogenesis. Plant Cell 1:747–755PubMedGoogle Scholar
  15. 15.
    Karr DB, Emerich DW (1987) Uniformity of the microsymbiont population from soybean nodules with respect to buoyant density. Plant Physiol 86:693–699Google Scholar
  16. 16.
    Klein S, Hirsch AM, Smith CA, Signer ER (1988) Interaction ofnod andexo Rhizobium meliloti in alfalfa nodulation. Mol Plant-Microbe Interact 1:94–100PubMedGoogle Scholar
  17. 17.
    Klein S, Walker GC, Signer ER (1988) All nod genes ofRhizobium meliloti are involved in alfalfa nodulation byexo mutants. J Bacteriol 170:1003–1006PubMedGoogle Scholar
  18. 18.
    Law, IJ, Yamamoto Y, Mort AJ, Bauer WD (1982) Nodulation of soybean byRhizobium japonicum mutants with altered capsule synthesis. Planta 154:100–109Google Scholar
  19. 19.
    Leigh JA, Lee CC (1988) Characterization of polysaccharides ofRhizobium meliloti exo mutants that form ineffective nodules. J Bacteriol 170:3327–3332PubMedGoogle Scholar
  20. 20.
    Leigh JA, Signer ER, Walker GC (1985) Exopolysaccharide-deficient mutants ofRhizobium meliloti that form ineffective nodules. Proc Natl Acad Sci USA 82:6231–6235PubMedGoogle Scholar
  21. 21.
    Leigh JA, Reed JW, Hanks JF, Hirsch AM, Walker GC (1987)Rhizobium meliloti mutants that fail to succinylate their calcifluor-binding exopolysaccharide are defective in nodule invasion. Cell 51:579–587PubMedGoogle Scholar
  22. 22.
    Long S, Reed JW, Himawan J, Walker GC (1988) Genetic analysis of a cluster of genes required for synthesis of the calcifluor-binding exopolysaccharide ofRhizobium meliloti. J Bacteriol 170:4239–4248PubMedGoogle Scholar
  23. 23.
    Mort AJ, Bauer WD (1982) Application of two new methods for cleavage of polysaccharides into specific oligosaccharide fragments: structure of the capsular and extracellular polysaccharides ofRhizobium japonicum that bind soybean lectin. J Biol Chem 257:1870–1875PubMedGoogle Scholar
  24. 24.
    Napoli C, Albersheim P (1980)Rhizobium leguminosarum mutants incapable of normal extracellular polysaccharide production. J Bacteriol 141:1454–1456PubMedGoogle Scholar
  25. 25.
    Nguyen LK, Schiller NL (1989) Identification of a slime exopolysaccharide depolymerase in mucoid strains ofPseudomonas aeruginosa. Curr Microbiol. 18:323–329Google Scholar
  26. 26.
    Olivares J, Bedmar EJ, Martinez-Molina E (1984) Infectivity ofRhizobium meliloti as affected by extracellular polysaccharides. J Appl Bacteriol 56:389–393Google Scholar
  27. 27.
    Preiss J, Ashwell G (1962) Alginic acid metabolism in bacteria. I. Enzymatic formation of unsaturated oligosaccharides and 4-deoxy-L-erythro-5-hexoseulose uronic acid. J Biol Chem 237:309–316PubMedGoogle Scholar
  28. 28.
    Sijam K, Goodman RN, Karr AL (1985) The effects of salts on the viscosity and wilt inducing capacity of the capsular polysaccharide ofErwinia amylovora. Physiol Plant Pathol. 26:231–239Google Scholar
  29. 29.
    Sutherland IW (1977) Enzymes acting on bacterial surface carbohydrates, In: Sutherland I (ed), Surface carbohydrates of the prokaryotic cell. New York: Academic Press, pp 209–245Google Scholar
  30. 30.
    Tully RE (1988) Synthesis of exopolysaccharide byBradyrhizobium japonicum during growth on hydroaromatic substrates. Appl Environ Microbiol 54:1624–1626Google Scholar
  31. 31.
    Zahn H, Levery SB, Lee CC, Leigh JA (1989) A second exopolysaccharide ofRhizobium meliloti strain SU47 that can function in root nodule invasion. Proc Natl Acad Sci USA. 86:3055–3059PubMedGoogle Scholar

Copyright information

© Springer-Verlag New York Inc. 1990

Authors and Affiliations

  • Michael F. Dunn
    • 1
  • Arthur L. Karr
    • 1
  1. 1.Department of Plant Pathology and the Interdisciplinary Plant GroupUniversity of MissouriColumbiaUSA

Personalised recommendations