Purification of a functional receptor for Clostridium difficile toxin A from intestinal brush border membranes of infant hamsters.
A receptor for Clostridium difficile toxin A was purified from brush border membranes (BBMs) from the small intestine of infant hamsters. The BBMs were solubilized with Triton X-114, and the solubilized receptor was purified with use of a toxin A immobilized affinity-chromatography column and differential temperature elution. SDS-PAGE and silver staining of the purified receptor revealed numerous high-molecular-weight bands. However, ligand blotting analysis with 125I-toxin A used as the probe identified a 163-kD protein as the predominate toxin A-binding molecule. Toxin A bound to the purified receptor at physiological temperature, but the amount of toxin bound increased at lower temperatures. Bovine thyroglobulin bound to toxin A and inhibited its binding to the purified receptor. Preincubation of the receptor with lectins produced by Bandeirea simplicifolia or Datura stramonium reduced specific binding by 125I-toxin A. Our data indicate that the purified toxin A receptor from small intestine BBMs of infant hamsters is a galactose- and N-acetylglucosamine-containing glycoprotein.
Rolfe
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RD
Department of Microbiology, School of Medicine, Texas Tech University Health Sciences Center, Lubbock 79430.
Song
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W
D000818:Animals; D001427:Bacterial Toxins; D001667:Binding, Competitive; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D016360:Clostridioides difficile / Q000378:metabolism; D006224:Cricetinae; D004768:Enterotoxins / Q000378:metabolism*; D005260:Female; D066298:In Vitro Techniques; D007421:Intestine, Small / Q000378:metabolism / Q000382:microbiology*; D037102:Lectins / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D008647:Mesocricetus; D008871:Microvilli / Q000378:metabolism / Q000382:microbiology; D008969:Molecular Sequence Data; D013696:Temperature; D013954:Thyroglobulin / Q000737:chemistry / Q000378:metabolism
D001427:Bacterial Toxins; D004768:Enterotoxins; D037102:Lectins; D008565:Membrane Proteins; C041348:tcdA protein, Clostridium difficile; D013954:Thyroglobulin