Abstract
Bacterial signal transduction systems commonly use receiver (REC) domains, which regulate adaptive responses to the environment as a function of their phosphorylation state. REC domains control cell physiology through diverse mechanisms, many of which remain understudied. We have defined structural features that underlie activation of the multi-domain REC protein, PhyR, which functions as an anti-anti-σ factor and regulates transcription of genes required for stress adaptation and host-microbe interactions in Alphaproteobacteria. Though REC phosphorylation is necessary for PhyR function in vivo, we did not detect expected changes in inter-domain interactions upon phosphorylation by solution X-ray scattering. We sought to understand this result by defining additional molecular requirements for PhyR activation. We uncovered specific interactions between unphosphorylated PhyR and an intrinsically disordered region (IDR) of the anti-σ factor, NepR, by solution NMR spectroscopy. Our data support a model whereby nascent NepR(IDR)-PhyR interactions and REC phosphorylation coordinately impart the free energy to shift PhyR to an open, active conformation that binds and inhibits NepR. This mechanism ensures PhyR is activated only when NepR and an activating phosphoryl signal are present. Our study provides new structural understanding of the molecular regulatory logic underlying a conserved environmental response system.