We have reconstructed the three-dimensional quaternary structure of the complete 480 kDa insulin receptor (IR), complexed with NanoGold-labelled insulin, via sets of electron micrographs obtained by low-dose low-temperature dark field scanning transmission electron microscopy (STEM).

Insulin binding to IR in mammalian cell membranes is essential for its manifold effects such as glucose homeostasis, increased protein synthesis, growth, and development. IR belongs to the superfamily of transmembrane receptor tyrosine kinases that include the monomeric epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor (PDGFR). In contrast, IR and its homologues IGF-1R (insulin-like growth factor 1 receptor) and IRR (insulin receptorrelated receptor) are sub-types of this family that are intrinsic disulfide-linked dimers of two αβ heterodimers. Monomeric receptor TKs are inactive, but are activated by ligand-induced dimerization that results in autophosphorylation. IR-like TKs are also inactive even though they are already dimeric, and are activated by ligand binding without further oligomerization. Insulin binding to the extracellular domain of IR results in autophosphorylation of specific tyrosines to initiate an intracellular signal transduction cascade. However, because the quaternary structure of IR is not known, the structural basis for the mechanism of IR activation by extracellular insulin binding has not been elucidated.

The insulin receptor was purified from human placenta. Bovine insulin was derivatized with NanoGold at the B chain Phel, a location not directly involved in receptor binding. Binding of derivatized insulin to the purified receptor was reduced only slightly compared to binding of the native insulin.