TLR2-stimulating contaminants in glycoconjugate fractions prepared from Bacteroides fragilis

Innate Immun. 2017 Jul;23(5):449-458. doi: 10.1177/1753425917714313. Epub 2017 Jun 13.


Bacteroides fragilis is a member of the normal intestinal flora and is involved in host immunostimulation via TLR2. On the bacterial cell surface, glycoconjugates, such as LPS and capsular polysaccharide A (PSA), have been reported to participate in host immunostimulation via TLR2. Previously, we identified a TLR2-stimulating lipoprotein in B. fragilis cells. In this study, we demonstrated that TLR2-stimulating principal molecules in glycoconjugate fractions prepared from B. fragilis are contaminating proteinous molecules, which may also be lipoproteins. The glycoconjugate fractions were prepared by phenol-hot water extraction of B. fragilis wild type and PSA-deficient strains, followed by hydrophobic interaction chromatography. TLR2-stimilating activities of the fractions were not affected by PSA deficiency. By in-gel TLR2-stimulation assay, molecules in high-molecular-mass area, where capsular polysaccharides were migrated, were found not to stimulate TLR2, but those in the range of 15-40 kDa were active. Further, proteinase K could digest the latter molecules and the TLR2-stimulating activities were migrated to the area of below 15 kDa. These results support that proteinous molecules, which are estimated to be lipoproteins, are responsible for almost all TLR2-stimulating activity in the glycoconjugate fractions prepared from B. fragilis.

Keywords: Bacteroides fragilis; Toll-like receptor; capsular polysaccharide; lipopolysaccharide; lipoprotein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Outer Membrane Proteins / genetics
  • Bacterial Outer Membrane Proteins / immunology
  • Bacterial Outer Membrane Proteins / metabolism*
  • Bacteroides Infections / immunology*
  • Bacteroides Infections / microbiology
  • Bacteroides fragilis / genetics
  • Bacteroides fragilis / immunology
  • Bacteroides fragilis / metabolism*
  • Cell Fractionation
  • Glycoconjugates / genetics
  • Glycoconjugates / metabolism*
  • Humans
  • Intestines / immunology*
  • Intestines / microbiology
  • Lipoproteins / genetics
  • Lipoproteins / immunology
  • Lipoproteins / metabolism*
  • Microorganisms, Genetically-Modified
  • Toll-Like Receptor 2 / metabolism*


  • Bacterial Outer Membrane Proteins
  • Glycoconjugates
  • Lipoproteins
  • TLR2 protein, human
  • Toll-Like Receptor 2