Yunn, N.-O. | Search bioRxiv

bioRxiv2025

Structural mechanism of insulin receptor activation by a dimeric aptamer agonist

Insulin binding to the insulin receptor (IR) triggers signaling pathways that regulate glucose uptake and cell growth. In previous work, we identified a DNA aptamer, A62, which partially activates the IR. During engineering aptamers for improved in vivo stability, we discovered that crosslinking two A62 aptamers with linkers of varying lengths led to full phosphorylation of the IR, though activation remained selective to the AKT pathway. To elucidate the mechanism behind this aptamer-induced full activation of the IR, we determined the structure of the IR in complex with a dimeric form of A62 (A62D) linked by an 8-nucleotide connector. We identified three distinct conformations of the IR: arrowhead-shaped, pseudo-arrowhead-shaped, and pseudo-gamma-shaped. The pseudo-gamma-shaped conformation closely resembles the structure of a fully active IR bound by a single insulin molecule. In these configurations, only one A62 monomer (A62M) within the A62D dimer binds to the IR dimer. This binding brings the IR monomers into close proximity, promoting intermolecular trans-phosphorylation. Our findings provide valuable structural insights for the development of novel therapeutic strategies targeting the IR.

biochemistry↗

bioRxiv2019

Stepwise autophosphorylation regulates biased agonism of the insulin receptor

Insulin is a key regulator of energy metabolism in peripheral tissues but also functions as a growth factor. Insulin binding to the insulin receptor (IR) leads to autophosphorylation of intracellular tyrosine residues, which simultaneously initiates a multitude of signals and functions. In contrast, some artificial (non-insulin) ligands for the IR result in biased agonism, selectively activating the PI3K/AKT pathway and metabolic effects without activating mitogen-activated protein kinase (MAPK) pathway and mitogenic effects. However, the precise mechanism of biased agonism at the receptor level remains unclear. The biased agonist IR-A48 aptamer selectively induces mono-phosphorylation of Tyr1150 residue (m-pY1150) in the kinase domain of IR. Hence, we hypothesized that IR autophosphorylation is a stepwise process in which formation of m-pY1150 represents an intermediate step. To explore this idea, we used hybrid receptors in which insulin can bind only one of the two binding sites of the dimeric receptor. Asymmetric insulin binding selectively induced symmetric m-pY1150 in both kinase domains. Moreover, the juxtamembrane domain, which interacts with the kinase domain, restricted the full activation of IR, and symmetric m-pY1150 played a crucial role in the rearrangement of intracellular domains to release this restriction. Our findings demonstrate that the symmetry of insulin binding to a dimeric receptor determines the stepwise autophosphorylation of IR. Furthermore, considering that the degree of ligand symmetry on a receptor depends mainly on ligand concentration, our results suggest that IR may play metabolic-biased roles in peripheral tissues dependent on local insulin concentrations.

cell biology↗