Synthetic pentapeptides inhibiting autophosphorylation of insulin receptor in a non-ATP-competitive mechanism

In an attempt to develop non-ATP-competitive inhibitors of the autophosphorylation of IR, the effects of the synthetic peptides, Ac-DIY¹¹⁵⁸ET-NH₂ and Ac-DY¹¹⁶²Y^1163RK-NH₂, on the phosphorylation of IR were studied in vitro. The peptides were derived from the amino-acid sequence in the activation loop of IR. They inhibited the autophosphorylation of IR to 20.5 and 40.7%, respectively, at 4000 μM. The Asp/Asn- and Glu/Gln-substituted peptides, Ac-NIYQT-NH₂ and Ac-NYYRK-NH₂, more potently inhibited the autophosphorylation than did the corresponding parent peptides. The inhibitory potencies of the substituted peptides were decreased with increasing concentrations of ATP, indicating that these peptides employ an ATP-competitive mechanism in inhibiting the autophosphorylation of IR. In contrast, those of the parent peptides were not affected. Mass spectrometry showed that the parent peptides were phosphorylated by IR, suggesting that they interact with the catalytic loop. Moreover, docking simulations predicted that the substituted peptides would interact with the ATP-binding region of IR, whereas their parent peptides would interact with the catalytic loop of IR. Thus, Ac-DIYET-NH₂ and Ac-DYYRK-NH₂ are expected to be non-ATP-competitive inhibitors. These peptides could contribute to the development of a drug employing a novel mechanism.