Dissociation of oxygen from the heme domain of the bacterial oxygen sensor protein FixL constitutes the first step in hypoxiainduced signaling. In the present study, the photodissociation of the heme-O2 bond was used to synchronize this event, and timeresolved resonance Raman (TR 3 ) spectroscopy with subpicosecond time resolution was implemented to characterize the heme configuration of the primary photoproduct. TR 3 measurements on heme-oxycomplexes are highly challenging and have not yet been reported. Whereas in all other known six-coordinated heme protein complexes with diatomic ligands, including the oxymyoglobin reported here, heme iron out-of-plane motion (doming) occurs faster than 1 ps after iron-ligand bond breaking; surprisingly, no sizeable doming is observed in the oxycomplex of the Bradyrhizobium japonicum FixL sensor domain (FixLH). This assessment is deduced from the absence of the iron-histidine band around 217 cm ؊1 as early as 0.5 ps. We suggest that efficient ultrafast oxygen rebinding to the heme occurs on the femtosecond time scale, thus hindering heme doming. Comparing WT oxy-FixLH, mutant proteins FixLH-R220H and FixLH-R220Q, the respective carbonmonoxy-complexes, and oxymyoglobin, we show that a hydrogen bond of the terminal oxygen atom with the residue in position 220 is responsible for the observed behavior; in WT FixL this residue is arginine, crucially implicated in signal transmission. We propose that the rigid O2 configuration imposed by this residue, in combination with the hydrophobic and constrained properties of the distal cavity, keep dissociated oxygen in place. These results uncover the origin of the ''oxygen cage'' properties of this oxygen sensor protein.heme protein ͉ molecular dynamics ͉ ultrafast spectroscopy ͉ vibrational spectroscopy

Background: DNR is a recently discovered transcriptional regulator homologous to the CO sensor CooA. Results: NO binding to heme selectively allows activation; ligand dynamics energetics implies common mechanism for 6-coordinate heme proteins. Conclusion: DNR is an NO sensor acting as ligand trap. Significance: Demonstration of NO-sensing function helps unravel signaling in the pathogen P. aeruginosa and describes common mechanism in emerging class of 6-coordinate heme proteins.

We investigated the removal of nitrogen and phosphate from the effluent of a sewage treatment plant over a long-term operation in bioreactors packed with different combinations of wood and iron, with a trickling filter packed with foam ceramics for nitrification. The average nitrification rate in the trickling filter was 0.17 kg N/m3∙day and remained at 0.11 kg N/m3∙day even when the water temperature was below 15 °C. The denitrification and phosphate removal rates in the bioreactor packed with aspen wood and iron were higher than those in the bioreactor packed with cedar chips and iron. The bioreactor packed with aspen wood and iron continued to remove nitrate and phosphate for >1200 days of operation. The nitrate removal activity of a biofilm attached to the aspen wood from the bioreactor after 784 days of operation was 0.42 g NO3-N/kg dry weight wood∙ day. There was no increase in the amount of dissolved organic matter in the outflow from the bioreactors.