α-Helical element at the hormone-binding surface of the insulin receptor functions as a signaling element to activate its tyrosine kinase.

The primary hormone-binding surface of the insulin receptor spans one face of the N-terminal ß-helix of the cc-subunit (the L1 domain) and an a-helix in its C-terminal segment (aCT). Crystallographic analysis of the free ectodomain has defined a contiguous dimer-related motif in which the aCT a-helix packs against L1 ß-strands 2 and 3. To relate structure to function, we exploited expanded genetic-code technology to insert photo-activatable probes at key sites in LI and aCT. The pattern of aCT-mediated photo-cross-linking within the free and bound receptor is in accord with the crystal structure and prior mutagenesis. Surprisingly, L1 photo-probes in ß-strands 2 and 3, predicted to be shielded by aCT, efficiently cross-link to insulin. Furthermore, anomalous mutations were identified on neighboring surfaces of aCT and insulin that impair hormone-dependent activation of the intracellular receptor tyrosine kinase (contained within the transmembrane ß-subunit) disproportionately to their effects on insulin binding. Taken together, these results suggest that aCT, in addition to its hormone-recognition role, provides a signaling element in the mechanism of receptor activation. [ABSTRACT FROM AUTHOR]