Binding specificity of Salmonella plasmid-encoded fimbriae assessed by glycomics.
The Salmonella enterica serotype Typhimurium (S. Typhimurium) genome encodes 12 intestinal colonization factors of the chaperone/usher fimbrial assembly class; however, the binding specificity is known for only one of these adhesins, known as type 1 fimbriae. Here we explored the utility of glycomics to determine the carbohydrate binding specificity of plasmid-encoded fimbriae from S. Typhimurium. A cosmid carrying the pef operon was introduced into Escherichia coli and expression of fimbrial filaments composed of PefA confirmed by flow cytometry and immune-electron microscopy. Plasmid-encoded fimbriae were purified from the surface of E. coli, and the resulting preparation was shown to contain PefA as the sole major protein component. The binding of purified plasmid-encoded fimbriae to a glycanarray suggested that this adhesin specifically binds the trisaccharide Galbeta1-4(Fucalpha1-3)GlcNAc, also known as the Lewis X (Le(x)) blood group antigen. Results from the glycanarray were validated by enzyme-linked immunosorbent assay (ELISA) in which plasmid-encoded fimbriae bound Le(x)-coated wells in a concentration-dependent manner. The binding of plasmid-encoded fimbriae to Le(x)-coated wells could be inhibited by co-incubation with soluble Le(x) antigen. Our results establish glycomic analysis as a promising new approach for determining the carbohydrate binding specificity of bacterial adhesins.
Chessa
Daniela
D
Department of Medical Microbiology and Immunology, School of Medicine, University of California at Davis, Davis, CA 95616-8645, USA.
Dorsey
Caleb W
CW
Winter
Maria
M
Baümler
Andreas J
AJ
D000595:Amino Acid Sequence; D050260:Carbohydrate Metabolism; D036781:Fimbriae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010861:Fimbriae, Bacterial / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism* / Q000648:ultrastructure; D015964:Gene Expression Regulation, Bacterial; D054794:Glycomics; D008024:Ligands; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics* / Q000302:isolation & purification; D012475:Salmonella / Q000235:genetics* / Q000378:metabolism*
D008024:Ligands; D036781:Fimbriae Proteins