Topology of the hog intrinsic factor receptor in the intestine.
The intrinsic factor receptor was isolated from Triton X-100 extract of hog ileal mucosa using affinity chromatography on intrinsic factor bound to cobalamin-Sepharose. We verified that the receptor contains two subunits, alpha and beta. The purified receptor located in the detergent micelle was radioiodinated. The alpha subunit was labeled and dissociated from the receptor. When the receptor was immobilized on intrinsic factor cobalamin-Sepharose, the part of the receptor which binds intrinsic factor evidently faces the gel and the rest faces outward. When such gel-bound pure receptor was iodinated, the beta subunit was labeled. Iodination of the micellar cobalamin-intrinsic factor receptor complex also caused labeling of the beta subunit. This was interpreted as being due to a conformational change in the receptor affected by the binding of the substrate cobalamin-intrinsic factor, exposing groups accessible to iodination. The beta subunit was found to be hydrophobic, but the alpha subunit was soluble in phosphate buffer without detergent. The receptor was liberated from intestinal mucosa by papain treatment. The enzyme seems to solubilize an intrinsic factor-binding part of the receptor, apparently a part of the alpha subunit. The liberated papain-alpha was purified by affinity chromatography. In gel filtration, it seemed to occur in dimeric form, but its true Mr = 45,000, according to findings in sodium dodecyl sulfate-electrophoresis. In the light of the findings, the topology of the receptor is suggested to be as follows: the alpha subunit binding intrinsic factor faces out and the hydrophobic beta subunit faces in.
D000818:Animals; D007082:Ileum / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000378:metabolism*; D007437:Intrinsic Factor / Q000378:metabolism*; D046911:Macromolecular Substances; D008961:Models, Structural; D008970:Molecular Weight; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000302:isolation & purification* / Q000378:metabolism; D018000:Receptors, Peptide; D013552:Swine
D046911:Macromolecular Substances; D011956:Receptors, Cell Surface; D018000:Receptors, Peptide; C028199:intrinsic factor receptor; D007437:Intrinsic Factor