Dimerization Promotes\nPKR Activation by Modulating\nEnergetics of αC Helix Conversion between Active and Inactive\nConformations

Protein kinase R (PKR) functions in the eukaryotic innate\nimmune\nsystem as a first-line defense against viral infections. PKR binds\nviral dsRNA, leading to autophosphorylation and activation. In its\nactive state, PKR can phosphorylate its primary substrate, eIF2α,\nwhich blocks the initiation of translation in the infected cell. It\nhas been established that PKR activation occurs when the kinase domain\ndimerizes in a back-to-back configuration. However, the mechanism\nby which dimerization leads to enzymatic activation is not fully understood.\nHere, we investigate the structural mechanistic basis and energy landscape\nfor PKR activation, with a focus on the αC helixa kinase\nactivation and signal integration hubusing all-atom equilibrium\nand enhanced sampling molecular dynamics simulations. By employing\nwindow-exchange umbrella sampling, we compute free-energy profiles\nof activation, which show that back-to-back dimerization stabilizes\na catalytically competent conformation of PKR. Key hydrophobic residues\nin the homodimer interface contribute to stabilization of the αC\nhelix in an active conformation and the position of its critical glutamate\nresidue. Using linear mutual information analysis, we analyze allosteric\ncommunication connecting the protomers’ N-lobes and the αC\nhelix dimer interface with the αC helix.