Solubilization of the ileal receptor intrinsic factor-vitamin B12 comples in the rat

Solubilization of the ileal receptor for intrinsic factor-B12 complex (IF-B12) was attempted by extracting ileal mucosa of the rat with an alkaline buffer of pH 10 and mechanical grinding. The ileal extract, when incubated with homologous IF-57CoB12 and applied on a bio-Gel A-5m column, produced a macromolecular fraction containing IF-57CoB12, which is presumed to be a complex of receptor (Rec) and IF-B12. Ileal extract after centrifugation at 100 000 g for 1 hour still yielded Rec-IF-B12. Formation of this complex in vitro was also demonstrated by agar-gel electrophoresis in which a new peak appeared near the origin. This fraction was in the void volume of a Sephadex G-200 column. Ileal extracts obtained 1.5 and 3 hours after oral administration of 57CoB12 contained Rec-IF-57CoB12. Treatment of Rec-IF-B12 with EDTA or trypsin released IF-B12, suggesting involvement of divalent cations in Rec-IF-B12 and a protein nature of Rec.