A PAS domain with an oxygen labile [4Fe-4S](2+) cluster in the oxygen sensor kinase NreB of Staphylococcus carnosus.
The cytoplasmic histidine sensor kinase NreB of Staphylococcus carnosus responds to O(2) and controls together with the response regulator NreC the expression of genes of nitrate/nitrite respiration. nreBC homologous genes were found in Staphylococcus strains and Bacillus clausii, and a modified form was found in some Lactobacillus strains. NreB contains a sensory domain with similarity to heme B binding PAS domains. Anaerobically prepared NreB of S. carnosus exhibited a (diamagnetic) [4Fe-4S](2+) cluster when assessed by Mossbauer spectroscopy. Upon reaction with air, the cluster was degraded with a half-life of approximately 2.5 min. No significant amounts of Mossbauer or EPR detectable intermediates were found during the decay, but magnetic Mossbauer spectra revealed formation of diamagnetic [2Fe-2S](2+) clusters. After extended exposure to air, NreB was devoid of a FeS cluster. Photoreduction with deazaflavin produced small amounts of [4Fe-4S](+), which were degraded subsequently. The magnetically perturbed Mossbauer spectrum of the [4Fe-4S](2+) cluster corroborated the S = 0 spin state and revealed uniform electric field gradient tensors of the iron sites, suggesting full delocalization of the valence electrons and binding of each of the Fe ions by four S ligands, including the ligand to the protein. Mutation of each of the four Cys residues inactivated NreB function in vivo in accordance with their role as ligands. [4Fe-4S](2+) cluster-containing NreB had high kinase activity. Exposure to air decreased the kinase activity and content of the [4Fe-4S](2+) cluster with similar half-lives. We conclude that the sensory domain of NreB represents a new type of PAS domain containing a [4Fe-4S](2+) cluster for sensing and function.
Müllner
Martin
M
Institut fur Mikrobiologie and Weinforschung, Universitat Mainz, Becherweg 15, 55099 Mainz, Germany.
Hammel
Oliver
O
Mienert
Bernd
B
Schlag
Steffen
S
Bill
Eckhard
E
Unden
Gottfried
G
D000388:Air; D006207:Half-Life; D000071677:Histidine Kinase; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D008280:Magnetics; D010100:Oxygen / Q000737:chemistry / Q000494:pharmacology*; D011494:Protein Kinases / Q000737:chemistry*; D015204:Spectroscopy, Mossbauer; D013210:Staphylococcus / Q000201:enzymology*
D007506:Iron-Sulfur Proteins; D011494:Protein Kinases; D000071677:Histidine Kinase; D010100:Oxygen