INSULIN-RECEPTORS ARE BIVALENT AS DEMONSTRATED BY PHOTOAFFINITY-LABELING

Insulin receptors in human placental membranes were photoaffinity-labeled with a radioactive human insulin-like growth factor I (hIGF-I) photoprobe N(epsilon-B28)-monoazidobenzoyl I-125-hIGF-I either alone or together with a non-radioactive insulin photoprobe N(epsilon-B29)-monoazidobenzoyl insulin. Precipitation of the solubilized receptors with anti-insulin antibody showed that receptors labeled with the radioactive hIGF-I photoprobe were detected in the immunoprecipitate only when photolabeling was carried out in the presence of the non-radioactive insulin photoprobe. Comparable results were obtained in converse experiments using a radioactive insulin photoprobe N(epsilon-B29)-monoazidobenzoyl I-125-insulin, a non-radioactive hIGF-I photoprobe N(epsilon-B28)-monoazidobenzoyl hIGF-I, and an antibody to hIGF-I. The amount of radioactive receptors precipitated by either the anti-insulin antibody or the anti-hIGHF-I antibody was close to the expected amount. These observations demonstrate that the insulin receptor is bivalent being capable of binding two molecules of ligand.