Purification and characterization of a receptor for the 987P pilus of Escherichia coli.
A receptor-containing fraction that caused visible aggregation of piliated Escherichia coli strain 987 bacteria was released into solution when brush borders from adult rabbit small intestines were stored. The receptor-containing fraction was separated into more than 50 bands by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, 4 of which contained carbohydrate as demonstrated by staining with periodic acid-Schiff. Receptor activity in the receptor-containing fraction was associated with a low-molecular-weight (less than 14,000) glycoprotein as measured by Western blots. Extraction of brush borders with chloroform-methanol-water (4:8:3) resulted in increased pilus 987P receptor activity in the aqueous extract. Gel filtration chromatography of sodium dodecyl sulfate-solubilized aqueous extract yielded a fraction with 987P receptor activity that contained only the low-molecular-weight glycoprotein. The purified 987P receptor contained 81% carbohydrate and 19% amino acids by weight. Isoelectric focusing separated the 987P receptor receptor into 3 peaks corresponding to isoelectric points of 2.2, 3.8, and 4.1. Each of the peaks contained 987P receptor activity. The 987P receptor activity was sensitive to periodate oxidation and to digestion with pronase. The strain 987-agglutinating activity of the receptor was inhibited by amino sugars and their N-acetylated derivatives, by compounds having a free amino group, by high concentrations of salt, and by lectins that recognize D-galactose or L-fucose.
Dean
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Isaacson
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D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001667:Binding, Competitive; D002241:Carbohydrates / Q000032:analysis; D004848:Epithelium / Q000382:microbiology; D004926:Escherichia coli / Q000502:physiology*; D010861:Fimbriae, Bacterial / Q000502:physiology*; D007421:Intestine, Small / Q000382:microbiology*; D007700:Kinetics; D008871:Microvilli / Q000382:microbiology; D008970:Molecular Weight; D011817:Rabbits; D011971:Receptors, Immunologic / Q000302:isolation & purification* / Q000502:physiology
D000596:Amino Acids; D002241:Carbohydrates; D011971:Receptors, Immunologic; C037872:pili, bacterial receptor