title: Cubilin P1297L mutation associated with hereditary megaloblastic anemia 1 causes impaired recognition of intrinsic factor-vitamin B(12) by cubilin.
abstract: Megaloblastic anemia 1 (MGA1) is an autosomal recessive disorder caused by the selective intestinal malabsorption of intrinsic factor (IF) and vitamin B(12)/cobalamin (Cbl) in complex. Most Finnish patients with MGA1 carry the disease-specific P1297L mutation (FM1) in the IF-B(12) receptor, cubilin. By site-directed mutagenesis, mammalian expression, and functional comparison of the purified wild-type and FM1 mutant forms of the IF-Cbl-binding cubilin region (CUB domains 5-8, amino acid 928-1386), we have investigated the functional implications of the P1297L mutation. Surface plasmon resonance analysis revealed that the P1297L substitution specifically increases the K(d) for IF-Cbl binding several-fold, largely by decreasing the association rate constant. In agreement with the binding data, the wild-type protein, but not the FM1 mutant protein, potently inhibits 37 degrees C uptake of iodine 125-IF-Cbl in cubilin-expressing epithelial cells. In conclusion, the data presented show a substantial loss in affinity of the FM1 mutant form of the IF-Cbl binding region of cubilin. This now explains the malabsorption of Cbl and Cbl-dependent anemia in MGA1 patients with the FM1 mutation. (Blood. 2000;96:405-409)
authors.affiliation: Department of Medical Biochemistry, University of Aarhus, Aarhus, Denmark.
authors.forename: M
authors.initials: M
authors.lastname: Kristiansen
authors.forename: M
authors.initials: M
authors.lastname: Aminoff
authors.forename: C
authors.initials: C
authors.lastname: Jacobsen
authors.forename: A
authors.initials: A
authors.lastname: de La Chapelle
authors.forename: R
authors.initials: R
authors.lastname: Krahe
authors.forename: P J
authors.initials: PJ
authors.lastname: Verroust
authors.forename: S K
authors.initials: SK
authors.lastname: Moestrup
mesh_terms: D000749:Anemia, Megaloblastic / Q000235:genetics*; D001665:Binding Sites; D003001:Cloning, Molecular; D015870:Gene Expression; D006801:Humans; D007437:Intrinsic Factor / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D016133:Polymerase Chain Reaction; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D013329:Structure-Activity Relationship; D020349:Surface Plasmon Resonance; D014162:Transfection; D014805:Vitamin B 12 / Q000378:metabolism*
chemical_list: D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; C029486:intrinsic factor-cobalamin receptor; D007437:Intrinsic Factor; D014805:Vitamin B 12