Skip to main content
NCBI home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1995 Jul;61(7):2577–2582. doi: 10.1128/aem.61.7.2577-2582.1995

Purification and Characterization of Conjugated Bile Salt Hydrolase from Bifidobacterium longum BB536

J Grill, F Schneider, J Crociani, J Ballongue
PMCID: PMC1388489  PMID: 16535071

Abstract

Bifidobacterium species deconjugate taurocholic, taurodeoxycholic, taurochenodeoxycholic, glycocholic, glycodeoxycholic, and glycochenodeoxycholic acids. The enzyme level increases in the growth phase. No increase in activity is observed for the cytoplasmic enzyme after addition of conjugated bile acids to a stationary-phase culture. Conjugated bile salt hydrolase (BSH) was purified from Bifidobacterium longum BB536. Its apparent molecular mass in denaturing polyacrylamide gel electrophoresis was ca. 40,000 Da. The intact enzyme had a relative molecular weight of ca. 250,000 as determined by gel filtration chromatography, suggesting that the native BSH of B. longum is probably a hexamer. The purified enzyme is active towards both glycine and taurine conjugates of cholate, deoxycholate, and chenodeoxycholate. The pH optimum is in the range of 5.5 to 6.5. A loss of BSH activity is observed after incubation at temperatures higher than 42(deg)C; at 60(deg)C, 50% of the BSH activity is lost. The importance of free sulfhydryl groups at the enzyme active center is suggested. For B. longum BB536, no significant difference in the initial rate of deconjugation and enzymatic efficiency appears between bile salts. The enzymatic efficiency is higher for B. longum BB536 than for other genera. In this paper, a new method which permits a display of BSH activity directly on polyacrylamide gels is described; this method confirms the molecular weight obtained for B. longum BB536 BSH.

Full Text

The Full Text of this article is available as a PDF (244.9 KB).

Selected References

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

  1. Batta A. K., Salen G., Shefer S. Transformation of bile acids into iso-bile acids by Clostridium perfringens: possible transport of 3 beta-hydrogen via the coenzyme. Hepatology. 1985 Nov-Dec;5(6):1126–1131. doi: 10.1002/hep.1840050611. [DOI] [PubMed] [Google Scholar]
  2. Benno Y., Sawada K., Mitsuoka T. The intestinal microflora of infants: composition of fecal flora in breast-fed and bottle-fed infants. Microbiol Immunol. 1984;28(9):975–986. doi: 10.1111/j.1348-0421.1984.tb00754.x. [DOI] [PubMed] [Google Scholar]
  3. Binder H. J., Filburn B., Floch M. Bile acid inhibition of intestinal anaerobic organisms. Am J Clin Nutr. 1975 Feb;28(2):119–125. doi: 10.1093/ajcn/28.2.119. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  5. Christiaens H., Leer R. J., Pouwels P. H., Verstraete W. Cloning and expression of a conjugated bile acid hydrolase gene from Lactobacillus plantarum by using a direct plate assay. Appl Environ Microbiol. 1992 Dec;58(12):3792–3798. doi: 10.1128/aem.58.12.3792-3798.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Church F. C., Porter D. H., Catignani G. L., Swaisgood H. E. An o-phthalaldehyde spectrophotometric assay for proteinases. Anal Biochem. 1985 May 1;146(2):343–348. doi: 10.1016/0003-2697(85)90549-4. [DOI] [PubMed] [Google Scholar]
  7. Feighner S. D., Dashkevicz M. P. Effect of dietary carbohydrates on bacterial cholyltaurine hydrolase in poultry intestinal homogenates. Appl Environ Microbiol. 1988 Feb;54(2):337–342. doi: 10.1128/aem.54.2.337-342.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Feighner S. D., Dashkevicz M. P. Subtherapeutic levels of antibiotics in poultry feeds and their effects on weight gain, feed efficiency, and bacterial cholyltaurine hydrolase activity. Appl Environ Microbiol. 1987 Feb;53(2):331–336. doi: 10.1128/aem.53.2.331-336.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ferrari A., Pacini N., Canzi E. A note on bile acids transformations by strains of Bifidobacterium. J Appl Bacteriol. 1980 Oct;49(2):193–197. doi: 10.1111/j.1365-2672.1980.tb05117.x. [DOI] [PubMed] [Google Scholar]
  10. Floch M. H., Gershengoren W., Diamond S., Hersh T. Cholic acid inhibition of intestinal bacteria. Am J Clin Nutr. 1970 Jan;23(1):8–10. doi: 10.1093/ajcn/23.1.8. [DOI] [PubMed] [Google Scholar]
  11. Floch M. H., Gershengoren W., Elliott S., Spiro H. M. Bile acid inhibition of the intestinal microflora--a function for simple bile acids? Gastroenterology. 1971 Aug;61(2):228–233. [PubMed] [Google Scholar]
  12. Gilliland S. E. Acidophilus milk products: a review of potential benefits to consumers. J Dairy Sci. 1989 Oct;72(10):2483–2494. doi: 10.3168/jds.S0022-0302(89)79389-9. [DOI] [PubMed] [Google Scholar]
  13. Gilliland S. E., Speck M. L. Deconjugation of bile acids by intestinal lactobacilli. Appl Environ Microbiol. 1977 Jan;33(1):15–18. doi: 10.1128/aem.33.1.15-18.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gopal-Srivastava R., Hylemon P. B. Purification and characterization of bile salt hydrolase from Clostridium perfringens. J Lipid Res. 1988 Aug;29(8):1079–1085. [PubMed] [Google Scholar]
  15. Gorbach S. L., Tabaqchali S. Bacteria, bile, and the small bowel. Gut. 1969 Dec;10(12):963–972. doi: 10.1136/gut.10.12.963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Huijghebaert S. M., Hofmann A. F. Influence of the amino acid moiety on deconjugation of bile acid amidates by cholylglycine hydrolase or human fecal cultures. J Lipid Res. 1986 Jul;27(7):742–752. [PubMed] [Google Scholar]
  17. Kawamoto K., Horibe I., Uchida K. Purification and characterization of a new hydrolase for conjugated bile acids, chenodeoxycholyltaurine hydrolase, from Bacteroides vulgatus. J Biochem. 1989 Dec;106(6):1049–1053. doi: 10.1093/oxfordjournals.jbchem.a122962. [DOI] [PubMed] [Google Scholar]
  18. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  19. Lundeen S. G., Savage D. C. Characterization and purification of bile salt hydrolase from Lactobacillus sp. strain 100-100. J Bacteriol. 1990 Aug;172(8):4171–4177. doi: 10.1128/jb.172.8.4171-4177.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lundeen S. G., Savage D. C. Characterization of an extracellular factor that stimulates bile salt hydrolase activity in Lactobacillus sp. strain 100-100. FEMS Microbiol Lett. 1992 Jul 1;73(1-2):121–126. doi: 10.1016/0378-1097(92)90594-e. [DOI] [PubMed] [Google Scholar]
  21. Lundeen S. G., Savage D. C. Multiple forms of bile salt hydrolase from Lactobacillus sp. strain 100-100. J Bacteriol. 1992 Nov;174(22):7217–7220. doi: 10.1128/jb.174.22.7217-7220.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Macfarlane G. T., Hay S., Gibson G. R. Influence of mucin on glycosidase, protease and arylamidase activities of human gut bacteria grown in a 3-stage continuous culture system. J Appl Bacteriol. 1989 May;66(5):407–417. doi: 10.1111/j.1365-2672.1989.tb05110.x. [DOI] [PubMed] [Google Scholar]
  23. Masuda N. Deconjugation of bile salts by Bacteroids and Clostridium. Microbiol Immunol. 1981;25(1):1–11. doi: 10.1111/j.1348-0421.1981.tb00001.x. [DOI] [PubMed] [Google Scholar]
  24. Midtvedt T., Norman A. Bile acid transformations by microbial strains belonging to genera found in intestinal contents. Acta Pathol Microbiol Scand. 1967;71(4):629–638. doi: 10.1111/j.1699-0463.1967.tb05183.x. [DOI] [PubMed] [Google Scholar]
  25. Midtvedt T., Norman A. Parameters in 7-alpha-dehydroxylation of bile acids by anaerobic lactobacilli. Acta Pathol Microbiol Scand. 1968;72(2):313–329. doi: 10.1111/j.1699-0463.1968.tb01345.x. [DOI] [PubMed] [Google Scholar]
  26. Nair P. P., Gordon M., Reback J. The enzymatic cleavage of the carbon-nitrogen bond in 3-alpha, 7-alpha, 12-alpha-trihydroxy-5-beta-cholan-24-oylglycine. J Biol Chem. 1967 Jan 10;242(1):7–11. [PubMed] [Google Scholar]
  27. Northfield T. C., Drasar B. S., Wright J. T. Value of small intestinal bile acid analysis in the diagnosis of the stagnant loop syndrome. Gut. 1973 May;14(5):341–347. [PMC free article] [PubMed] [Google Scholar]
  28. Savage D. C. Microbial ecology of the gastrointestinal tract. Annu Rev Microbiol. 1977;31:107–133. doi: 10.1146/annurev.mi.31.100177.000543. [DOI] [PubMed] [Google Scholar]
  29. Stellwag E. J., Hylemon P. B. Purification and characterization of bile salt hydrolase from Bacteroides fragilis subsp. fragilis. Biochim Biophys Acta. 1976 Nov 8;452(1):165–176. doi: 10.1016/0005-2744(76)90068-1. [DOI] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES