Identification of determinants that confer ligand specificity on the insulin receptor

• Published in: The Journal of biological chemistry Vol. 267; no. 19; pp. 13681 - 13686
• Main Authors: Andersen, A S, Kjeldsen, T, Wiberg, F C, Vissing, H, Schäffer, L, Rasmussen, J S, De Meyts, P, Møller, N P
• Format: Journal Article
• Language: English
• Published: Bethesda, MD American Society for Biochemistry and Molecular Biology 05-07-1992
• Subjects: Amino Acid Sequence; amino acids; Biological and medical sciences; Cell receptors; Cell structures and functions; Cross-Linking Reagents; Fundamental and applied biological sciences. Psychology; Hormone receptors. Growth factor receptors. Cytokine receptors. Prostaglandin receptors; Humans; insulin; Insulin-Like Growth Factor I - metabolism; Ligands; man; membrane proteins; Molecular and cellular biology; Molecular Sequence Data; N-terminus; protein-tyrosine kinase; Receptor, Insulin - metabolism; receptors; Receptors, Cell Surface - metabolism; Receptors, Somatomedin; Recombinant Fusion Proteins - metabolism; requirements; specificity; Substrate Specificity; Human; Site specificity; Ligand binding; Radiolabelling; Somatomedin C; Substrate specificity; Insulin; Hormonal receptor
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)42267-3

We have previously shown, using truncated soluble recombinant receptors, that substituting the 62 N-terminal amino acids of
the alpha subunit from the insulin-like growth factor I receptor (IGFIR) with the corresponding 68 amino acids from the insulin
receptor (IR) results in a chimeric receptor with an approximately 200-fold increase in affinity for insulin and only a 5-fold
decrease in insulin-like growth factor I (IGFI) affinity (Kjeldsen, T., Andersen, A. S., Wiberg, F. C., Rasmussen, J. S.,
Schäffer, L., Balschmidt, P., Møller, K. B., and Møller, N. P. H. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 4404-4408). We
demonstrate that these 68 N-terminal amino acids of the IR also confer insulin affinity on the intact IGFI holoreceptor both
in the membrane-bound state and when solubilized by Triton X-100. Furthermore, this domain can be subdivided into two regions
(amino acids 1-27 and 28-68 of the IR alpha subunit) that, when replacing the corresponding IGFIR sequences, increases the
insulin affinity of truncated soluble receptor chimeras 8- and 20-fold, respectively, with only minor effects on the IGFI
affinity. Within the latter of these two regions, we found that amino acids 38-68 of the IR, representing 13 amino acid differences
from IGFIR, confer the same 20-fold increase in insulin affinity on the IGFIR. Finally, the amino acids from position 42 to
50 are not responsible for this increase in insulin affinity. We thus propose that at least two determinants within the 68
N-terminal amino acids of the insulin receptor are involved in defining the ligand specificity of the insulin receptor, and
that one or a combination of the remaining seven amino acid differences between position 38 and 68 are involved in conferring
insulin affinity on the insulin receptor.