Structural resolution of a tandem hormone-binding element in the insulin receptor and its implications for design of peptide agonists

摘要

The C-terminal segment of the human insulin receptor α-chain (designated αCT) is critical to insulin binding as has been previously demonstrated by alanine scanning mutagenesis and photo-cross-linking. To date no information regarding the structure of this segment within the receptor has been available. We employ here the technique of thermal-factor sharpening to enhance the interpretability of the electron-density maps associated with the earlier crystal structure of the human insulin receptor ectodomain. The αCT segment is now resolved as being engaged with the central β-sheet of the first leucine-rich repeat (L1) domain of the receptor. The segment is α-helical in conformation and extends 11 residues N-terminal of the classical αCT segment boundary originally defined by peptide mapping. This tandem structural element (αCT-L1) thus defines the intact primary insulin-binding surface of the apo-receptor. The structure, together with isothermal titration calorimetry data of mutant αCT peptides binding to an insulin minireceptor, leads to the conclusion that putative “insulin-mimetic” peptides in the literature act at least in part as mimics of the αCT segment as well as of insulin. Photo-cross-linking by novel bifunctional insulin derivatives demonstrates that the interaction of insulin with the αCT segment and the L1 domain occurs in trans, i.e., these components of the primary binding site are contributed by alternate α-chains within the insulin receptor homodimer. The tandem structural element defines a new target for the design of insulin agonists for the treatment of diabetes mellitus.