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

作者：Whittaker Jonathan; Whittaker Linda J; Roberts Charles T Jr; Phillips Nelson B; Ismail Beigi Faramarz; Lawrencee Michael C; Weiss Michael A

来源：Proceedings of the National Academy of Sciences, 2012, 109(28): 11166-11171.

DOI：10.1073/pnas.1205681109

• nonstandard mutagenesis
• alanine scanning
• affinity

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

• 出版日期2012-7-10
• 单位河北医科大学