Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1985 Feb;161(2):493–499. doi: 10.1128/jb.161.2.493-499.1985

Location and characteristics of enzymes involved in the breakdown of polygalacturonic acid by Bacteroides thetaiotaomicron.

R E McCarthy, S F Kotarski, A A Salyers
PMCID: PMC214909  PMID: 3968032

Abstract

When Bacteroides thetaiotaomicron is grown in medium which contains polygalacturonic acid (PGA) as the sole carbon source, two different polygalacturonases are produced: a PGA lyase (EC 4.2.2.2) and a PGA hydrolase (EC 3.2.1.15). Both enzymes are cell associated. The PGA hydrolase appears to be an inner membrane protein. The PGA lyase is a soluble protein that associates with membranes under certain conditions. The PGA lyase was purified to apparent homogeneity. It has a molecular weight (from sodium dodecyl sulfate-polyacrylamide gel electrophoresis) of 74,000, a pH optimum of 8.7, a pI of 7.5, and a Km for PGA of 40 to 70 micrograms/ml. It requires calcium for maximal activity. The main product of this enzyme appears to be a disaccharide that contains a delta 4,5-unsaturated galacturonic acid residue. The PGA hydrolase can be solubilized from membranes with 2% Triton X-100 and has been partially purified. It has a pH optimum of 5.4 to 5.5, a pI of 4.7 to 4.9, and a Km for PGA of 350 to 400 micrograms/ml. The main product of this enzyme appears to be galacturonic acid. The specific activities of both PGA hydrolase and PGA lyase increase at the same rate when bacteria are exposed to PGA. The two enzymes therefore appear to be similarly regulated.

Full text

PDF
493

Images in this article

497Image
on p.497

Selected References

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

  1. DISCHE Z. New color reactions for determination of sugars in polysaccharides. Methods Biochem Anal. 1955;2:313–358. doi: 10.1002/9780470110188.ch11. [DOI] [PubMed] [Google Scholar]
  2. Davé B. A., Vaughn R. H. Purification and properties of an polygalacturonic acid trans-eliminase produced by Bacillus pumilus. J Bacteriol. 1971 Oct;108(1):166–174. doi: 10.1128/jb.108.1.166-174.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dygert S., Li L. H., Florida D., Thoma J. A. Determination of reducing sugar with improved precision. Anal Biochem. 1965 Dec;13(3):367–374. doi: 10.1016/0003-2697(65)90327-1. [DOI] [PubMed] [Google Scholar]
  4. GAREN A., LEVINTHAL C. A fine-structure genetic and chemical study of the enzyme alkaline phosphatase of E. coli. I. Purification and characterization of alkaline phosphatase. Biochim Biophys Acta. 1960 Mar 11;38:470–483. doi: 10.1016/0006-3002(60)91282-8. [DOI] [PubMed] [Google Scholar]
  5. Kotarski S. F., Salyers A. A. Effect of long generation times on growth of Bacteroides thetaiotaomicron in carbohydrate-induced continuous culture. J Bacteriol. 1981 Jun;146(3):853–860. doi: 10.1128/jb.146.3.853-860.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kotarski S. F., Salyers A. A. Isolation and characterization of outer membranes of Bacteroides thetaiotaomicron grown on different carbohydrates. J Bacteriol. 1984 Apr;158(1):102–109. doi: 10.1128/jb.158.1.102-109.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. MACMILLAN J. D., VAUGHN R. H. PURIFICATION AND PROPERTIES OF A POLYGALACTURONIC ACID-TRANS-ELIMINASE PRODUCED BY CLOSTRIDIUM MULTIFERMENTANS. Biochemistry. 1964 Apr;3:564–572. doi: 10.1021/bi00892a016. [DOI] [PubMed] [Google Scholar]
  9. Miller L., Macmillan J. D. Mode of action of pectic enzymes. II. Further purification of exopolygalacturonate lyase and pectinesterase from Clostridium multifermentans. J Bacteriol. 1970 Apr;102(1):72–78. doi: 10.1128/jb.102.1.72-78.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. NAGEL C. W., VAUGHN R. H. The characteristics of a polygalacturonase produced by Bacillus polymyxa. Arch Biochem Biophys. 1961 May;93:344–352. doi: 10.1016/0003-9861(61)90277-6. [DOI] [PubMed] [Google Scholar]
  11. Nagel C. W., Wilson T. M. Pectic acid lyases of Bacillus polymyxa. Appl Microbiol. 1970 Sep;20(3):374–383. doi: 10.1128/am.20.3.374-383.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Nasuno S., Starr M. P. Polygalacturonase of Erwinia carotovora. J Biol Chem. 1966 Nov 25;241(22):5298–5306. [PubMed] [Google Scholar]
  13. PREISS J., ASHWELL G. Polygalacturonic acid metabolism in bacteria. II. Formation and metabolism of 3-deoxy-D-glycero-2, 5-hexodiulosonic acid. J Biol Chem. 1963 May;238:1577–1583. [PubMed] [Google Scholar]
  14. Salyers A. A., Kotarski S. F. Induction of chondroitin sulfate lyase activity in Bacteroides thetaiotaomicron. J Bacteriol. 1980 Aug;143(2):781–788. doi: 10.1128/jb.143.2.781-788.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Salyers A. A., O'Brien M. Cellular location of enzymes involved in chondroitin sulfate breakdown by Bacteroides thetaiotaomicron. J Bacteriol. 1980 Aug;143(2):772–780. doi: 10.1128/jb.143.2.772-780.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Salyers A. A., Vercellotti J. R., West S. E., Wilkins T. D. Fermentation of mucin and plant polysaccharides by strains of Bacteroides from the human colon. Appl Environ Microbiol. 1977 Feb;33(2):319–322. doi: 10.1128/aem.33.2.319-322.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Varel V. H., Bryant M. P. Nutritional features of Bacteroides fragilis subsp. fragilis. Appl Microbiol. 1974 Aug;28(2):251–257. doi: 10.1128/am.28.2.251-257.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Wojciechowicz M., Heinrichova K., Ziołecki A. A polygalacturonate lyase produced by Lachnospira multiparus isolated from the bovine rumen. J Gen Microbiol. 1980 Mar;117(1):193–199. doi: 10.1099/00221287-117-1-193. [DOI] [PubMed] [Google Scholar]
  19. Wojciechowicz M. Partial characterization of pectinolytic enzymes of Bacteroides ruminicola isolated from the rumen of a sheep. Acta Microbiol Pol A. 1971;3(1):45–56. [PubMed] [Google Scholar]
  20. Wojciechowicz M., Ziołecki A. Pectinolytic enzymes of large rumen treponemes. Appl Environ Microbiol. 1979 Jan;37(1):136–142. doi: 10.1128/aem.37.1.136-142.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES