We have previously demonstrated that isolated insulin and IGF-1 αβ half-receptors can be reconstituted into a functional α2β2 hybrid receptor complex [Treadway et al. (1989) J. Biol. Chem. 264, 21450-21453]. In the present study, we have examined this assembly process by determining the effect of ligand occupancy and Mn/MgATP binding on the dimerization of mutant and wild-type insulin and IGF-1 a0 half-receptors. IGF-1 or Mn/MgAMPPCP binding to wild-type IGF-1 αβ half-receptors resulted in the specific assembly of the αβ half-receptors into an α2β2 heterotetrameric IGF-1 holoreceptor complex. Similarly, insulin binding to the kinase-deficient mutant (A/Kiois) insulin αβ half-receptor also resulted in the specific assembly into an 2/82 holoreceptor complex. In contrast, Mn/MgAMPPCP treatment of A/K1018 mutant insulin αβ half-receptors did not induce heterotetramer assembly, consistent with the inability of this mutant receptor to bind ATP. The ability of the insulin αβ receptors to assemble with the IGF-1 αβ half-receptors was used to examine the intermolecular subunit interactions responsible for dimerization. In the presence of Mn/MgAMPPCP, the wild-type insulin and wild-type IGF-1 aft half-receptors were observed to assemble into an insulin/IGF-1 α2β2 hybrid receptor complex. Similarly, a combination of insulin and IGF-1 induced hybrid receptor formation between wild-type IGF-1 and A/K1018 mutant insulin αβ half-receptors. In contrast, mixing insulin-occupied A/Kiois mutant insulin αβ half-receptors with Mn/MgAMPPCP-occupied wild-type IGF-1 αβ half-receptors did not result in hybrid receptor formation. Thus, these data demonstrate that ligand binding to the a subunit and Mn/MgATP binding to the α subunit do not result in complementary conformational changes necessary for αβ half-receptor dimerization.
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