Figure 1. PhyR α11-β5 loop rearrangement decreases the energetic barrier to opening.

(A) A model of general stress response (GSR) activation. Upstream sensory kinases phosphorylate PhyR, leading to PhyR opening and NepR binding. This releases σ^EcfG to direct GSR transcription. (B) Alignment of PhyR (green) and PhyR~P (gold) MD trajectories. Rearrangement can occur on short MD timescales in the α11-β5 loop, which retracts toward the REC domain (shown by arrow). Structures shown are the average backbone positions over the final 10 ns of simulation. (C) NepR (pink) modelled into the PhyR-SL domain (green) binding site using a hypothetical open state model. Hydrophobic (red) and hydrophilic (gray) regions at the domain interface (brown) are highlighted. (D) Calculated solvent accessible surface area (SASA) values for PhyR (green) and PhyR~P (gold) at 50 ns for NepR-binding residues. (E) Umbrella sampling of PhyR and PhyR~P at increasing inter-domain distances. Arrows indicate points corresponding to the initial equilibrium state of the protein, collected from previous simulations, as well as the point in the PhyR~P (50 ns) sampling where solvation of the total interdomain interface began. The energetic barrier to open PhyR is lowered after phosphorylation (see 50 ns simulation). Energy to open PhyR~P prior to solvation is equivalent to unphosphorylated PhyR. Error bars represent standard deviation.