Conformational changes induced by phosphorylation of the FixJ receiver domain

Background: A variety of bacterial adaptative cellular responses to environ mental stimuli are mediated by two-component signal transduction pathways. In these phosphorelay cascades, histidine kinases transphosphorylate a cons erved aspartate in the receiver domain, a conserved module in the response regulator superfamily. The main effect of this phosphorylation is to alter the conformation of the response regulator in order to modulate its biologi cal function. The response regulator FixJ displays a typical modular arrang ement, with a phosphorylatable N-terminal receiver domain and a C-terminal DNA-binding domain. In the symbiotic bacterium Sinorhizobium meliloti, phos phorylation of this response regulator activates transcription of nitrogen- fixation genes. Results: The crystal structures of the phosphorylated and of the unphosphor ylated N-terminal receiver domain of FixJ (FixJN) were solved at 2.3 Angstr om and 2.4 Angstrom resolution, respectively. They reveal the environment o f the phosphoaspartate in the active site and the specific conformational c hanges leading to activation of the response regulator, Phosphorylation of the conserved aspartate induces major structural changes in the beta 4-alph a 4 loop, and in the signaling surface alpha 4-beta 5 that mediates dimeriz ation of the phosphorylated full-length response regulator. A site-directed mutant at this protein-protein interface decreases the affinity of the pho sphorylated response regulator for the fixK promoter tenfold. Conclusions: The cascade of phosphorylation-induced conformational changes in FixJN illustrates the role of conserved residues in stabilizing the phos phoryl group in the active site, triggering the structural transition and a chieving the post-phosphorylation signaling events. We propose that these p hosphorylation-induced conformational changes underly the activation of res ponse regulators in general.