Abstract
In order to characterize regions of the insulin receptor that are essential for ligand binding we have minimized the ligand-binding domain of IR by site directed mutagenesis. The smallest receptor fragment identified comprised the first three domains of IR (1-468) fused to 16 amino acids from the C-terminal of the α-subunit The mass of this receptor fragment was 70 kD, and the affinity for insulin was 5 nM, which is similar to what was found for the full length soluble ectodomain of the insulin receptor. A similarly minimized IGF-I receptor construct (mIGF-IR) was shown to bind IGF-I with an affinity of 1.5 nM.
The role of the C-terminal peptide was further investigated by characterizing chimeric mini-receptor constructs. The carboxy terminal domain of the insulin receptor related receptor (IRRR) was found to abolish binding in IR and IGF-IR context, whereas swapping the carboxy terminal domains in either mIR or mIGF-IR context only cause minor changes in affinities, demonstrating that the carboxy terminal of IR and IGF-IR α-subunits are interchangable suggesting that this domain is part of the common binding site. The last part of this chapter describes a novel high affinity insulin binding protein (IBP) secreted from cells from at least three insects. This IBP is composed of two Ig-like C2-domains, has a molecular weight of 27 kDa, binds human insulin and related peptides with high affinity (10–200 pM) and inhibits insulin signaling through the insulin receptor. The ligand binding profile suggests that IBP recognizes a region that is highly conserved in the insulin superfamily but distinct from the classical insulin receptor-binding site.
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Kristensen, C., Andersen, A.S. (2002). Insulin Interaction with Minimized Receptors and Binding Proteins. In: Dieken, M.L., Federwisch, M., De Meyts, P. (eds) Insulin & Related Proteins - Structure to Function and Pharmacology. Springer, Dordrecht. https://doi.org/10.1007/0-306-47582-0_13
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DOI: https://doi.org/10.1007/0-306-47582-0_13
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