Ligand modulation of allosteric networks in an ancestral steroid receptor

Understanding the evolution of binding specificity, a heavily studied area of research, is key for determining how protein sequence changes alter function. Ligand-activation in the steroid receptor subfamily of transcription factors operates via a common allosteric mechanism which permits extant receptors to respond specifically to their cognate hormones. Here, we combine atomistic simulations with graph theory-based modeling of the inter-residue interactions within protein complexes to gain insight into how allostery drove selectivity in an ancestral receptor. An inactive ligand complex displays weakened allosteric communication, as quantified by suboptimal paths linking two functional surfaces. When function-switching mutations are incorporated, responses in allosteric networks are consistent with ligand activation status. Further analysis reveals residues that modulate features distinguishing active and inactive complexes, identifying a key, conserved residue that is crucial for activation in steroid receptors. We have identified a computational method using dynamic network analysis to probe the allosteric mechanisms driving the evolution of ligand specificity in hormone receptors, determining how residue substitutions altered allosteric networks to permit gain or loss of ligand response. These results may have general utility in elucidating how modern steroid receptors are activated by endogenous and xenobiotic molecules.\n\nAuthor summaryProteins interact with a host of biological partners to mediate their function. These binding partners are able to alter structural properties of the protein to send signals dictating downstream biological activity. This mode of regulation is described as allostery. Here, we perform a computational investigation of allostery in steroid receptors, a family of proteins that regulate a host of important biological processes in response to binding and activation by a steroidal ligand. We leverage a defined evolutionary system where known historical amino acid substitutions within the receptor drive a switch in ligand preference and receptor activation. We show that activating ligands induce stronger allosteric signaling between the ligand and the functional surface on the receptor. In addition, we incorporate evolutionary mutations that are known to alter ligand preference and show that this effect may be explained by allostery. This work provides insight into how amino acid substitutions over evolution affect allostery in proteins, permitting the loss and gain of function.