Transcriptional activation domains (ADs) are generally thought to be intrinsically unstructured, but capable of adopting limited secondary structure upon interaction with a coactivator surface. The indeterminate nature of this interface made it hitherto difficult to study structure/function relationships of such contacts. Here we used atomistic accelerated molecular dynamics (aMD) simulations to study the conformational changes of the GCN4 AD and variants thereof, either free in solution, or bound to the GAL11 coactivator surface. We show that the AD-coactivator interactions are highly dynamic while obeying distinct rules. The data provide insights into the constant and variable aspects of orientation of ADs relative to the coactivator, changes in secondary structure and energetic contributions stabilizing the various conformers at different time points. We also demonstrate that a prediction of α-helical propensity correlates directly with the experimentally measured transactivation potential of a large set of mutagenized ADs. The link between α-helical propensity and the stimulatory activity of ADs has fundamental practical and theoretical implications concerning the recruitment of ADs to coactivators.Author Summary

The regulated transcription of eukaryotic genes is governed by gene-specific transcription factors that contain activation domains to stimulate the expression of nearby genes. Activation domains are unable to take up a defined three-dimensional conformation. Nevertheless, as we demonstrate in our study, molecular dynamics simulations reveal that the key docking point of such domains (centered around several large hydrophobic amino acid sidechains) folds into fluctuating α-helical conformations. Analysis of published data shows that this tendency of adopting such local structures correlates directly with stimulation activity. We also investigate the interaction of these structurally unstable domains with a coactivator interaction partner. Computational simulations are ideally suited for analysing the rapidly changing, “fuzzy” interactions occurring between these protein partners. We gained new insights into the competitive nature of the key hydrophobic sidechains in binding to a pocket on the coactivator surface and documented for the first time the rapidly changing movements of an activation domain during these interactions.