FixL is a heme-based oxygen-sensing histidine kinase that induces expression of nitrogen fixation genes under hypoxic conditions. Oxygen binding to heme iron in the sensor domain of FixL initiates protein conformational changes that are transmitted to the histidine kinase domain, inactivating autophosphorylation activity. Although FixL also can bind other diatomic ligands such as CO, the CO-bound FixL represents incomplete inhibition of kinase activity. Ultraviolet resonance Raman (UVRR) spectra revealed that oxygen binding to the truncated sensor domain of FixL markedly decreased the intensity of the Y8a band arising from F-alpha-10 Tyr. In contrast, no appreciable change in intensity of the Y8a band occurred after CO binding, and time-resolved UVRR spectra of the sensor domain of FixL upon O-2 dissociation indicated that structural changes near F-alpha-10 Tyr occurred at similar to 0.1 mu s. These results suggest that O-2 dissociation from FixL changes the protein conformation near the F-alpha-10 Tyr residue within a microsecond. The conformational changes of FixL upon O-2 dissociation and the underlying sensing mechanism also are discussed.