Isolation and characterization of Bacteroides nodosus fimbriae: structural subunit and basal protein antigens.
We examined the isolation of fimbriae from Bacteroides nodosus. It was found that the best preparations were obtained from the supernatant of washed cells cultured on solid medium, from which fimbriae could be recovered in high yield and purity by a simple one-step procedure. Analysis of such preparations by sodium dodecyl sulfate gel electrophoresis showed that greater than 98% of the protein consisted of fimbrial structural subunits whose molecular weight was ca. 17,000. These preparations also usually exhibited minor contamination with a polypeptide of ca. 80,000 molecular weight, as well as trace amounts of lipopolysaccharide. Attempts to release additional fimbriae by the traditional means of subjecting the bacterial cells to physical stress, such as shearing or heating, resulted primarily in an increase in the level of contamination, without significant gain in the yield of fimbriae. Removal of the 80,000-dalton component could not be achieved by any of a variety of techniques normally used in fimbriae purification, including isoelectric precipitation, MgCl2 precipitation, and CsCl gradient ultracentrifugation, implying a direct physical association with the fimbrial strand. Electron micrographs of fractions containing this protein show cap-shaped structures attached to the ends of what appeared to be fimbrial stubs. These observations suggest that the 80,000-dalton polypeptide may actually constitute the basal attachment site which anchors the fimbria to the outer membrane, analogous to a similar protein recently described in enterotoxigenic strains of Escherichia coli. In B. nodosus, this 80,000-dalton protein is a major surface antigen, and like the fimbrial subunit, exhibited variation in electrophoretic mobility between serotypically different isolates.
D000942:Antigens, Bacterial / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000302:isolation & purification*; D001439:Bacteroides / Q000648:ultrastructure*; D004591:Electrophoresis, Polyacrylamide Gel; D010861:Fimbriae, Bacterial / Q000648:ultrastructure*; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008970:Molecular Weight
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D046911:Macromolecular Substances