title: High-affinity ligand binding by wild-type/mutant heteromeric complexes of the mannose 6-phosphate/insulin-like growth factor II receptor.
abstract: The mannose 6-phosphate/insulin-like growth factor II receptor has diverse ligand-binding properties contributing to its roles in lysosome biogenesis and growth suppression. Optimal receptor binding and internalization of mannose 6-phosphate (Man-6-P)-bearing ligands requires a dimeric structure leading to bivalent high-affinity binding, presumably mediated by cooperation between sites on both subunits. Insulin-like growth factor II (IGF-II) binds to a single site on each monomer. It is hypothesized that IGF-II binding to cognate sites on each monomer occurs independently, but bivalent Man-6-P ligand binding requires cooperative contributions from sites on both monomers. To test this hypothesis, we co-immunoprecipitated differentially epitope-tagged soluble mini-receptors and assessed ligand binding. Pairing of wild-type and point-mutated IGF-II binding sites between two dimerized mini-receptors had no effect on the function of the contralateral binding site, indicating IGF-II binding to each side of the dimer is independent and manifests no intersubunit effects. As expected, heterodimeric receptors composed of a wild-type monomer and a mutant bearing two Man-6-P-binding knockout mutations form functional IGF-II binding sites. By contrast to prediction, such heterodimeric receptors also bind Man-6-P-based ligands with high affinity, and the amount of binding can be attributed entirely to the immunoprecipitated wild-type receptors. Anchoring of both C-terminal ends of the heterodimer produces optimal binding of both IGF-II and Man-6-P ligands. Thus, IGF-II binds independently to both subunits of the dimeric mannose 6-phosphate/insulin-like growth factor II receptor. Although wild-type/mutant hetero-oligomers form readily when mixed, it appears that multivalent Man-6-P ligands bind preferentially to wild-type sites, possibly by cross-bridging receptors within clusters of immobilized receptors.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE 68198-5870, USA.
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Hartman
authors.forename: Jodi L
authors.initials: JL
authors.lastname: Kreiling
authors.forename: James C
authors.initials: JC
authors.lastname: Byrd
authors.forename: Richard G
authors.initials: RG
authors.lastname: MacDonald
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D019281:Dimerization; D006801:Humans; D047468:Immunoprecipitation; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D008360:Mannosephosphates / Q000378:metabolism*; D009154:Mutation; D017527:Receptor, IGF Type 2 / Q000235:genetics* / Q000378:metabolism*
chemical_list: D008024:Ligands; D008360:Mannosephosphates; D017527:Receptor, IGF Type 2; C027693:mannose-6-phosphate; D007335:Insulin-Like Growth Factor II