Functional implications of the propionate 7-arginine 220 interaction in the FixLH oxygen sensor from Bradyrhizobium japonicum

BjFixL from Bradyrhizobium japonicum is a heme-based oxygen sensor implicated in the signaling cascade that enables the bacterium to adapt to fluctuating oxygen levels. Signal transduction is initiated by the binding of O₂ to the heme domain of BjFixL, resulting in protein conformational changes that are transmitted to a histidine kinase domain. We report structural changes of the heme and its binding pocket in the Fe^II deoxy and Fe^III met states of the wild-type BjFixLH oxygen sensor domain and four mutants of the highly conserved residue arginine 220. UV-visible, electron paramagnetic resonance, and resonance Raman spectroscopies all showed that the heme iron of the R220H mutant is unexpectedly six-coordinated at physiological pH in the Fe^III state but undergoes pH- and redox-dependent coordination changes. This behavior is unprecedented for FixL proteins, but is reminiscent of another oxygen sensor from E. coli, EcDos. All mutants in their deoxy states are five-coordinated Fe^II, although we report rupture of the residue 220-propionate 7 interaction and structural modifications of the heme conformation as well as propionate geometry and flexibility. In this work, we conclude that part of the structural reorganization usually attributed to O₂ binding in the wild-type protein is in fact due to rupture of the Arg220-P7 interaction. Moreover, we correlate the structural modifications of the deoxy Fe^II states with _on values and conclude that the Arg220-P7 interaction is responsible for the lower O₂ and CO k _on values reported for the wild-type protein.