Table 1 Phosphotransfer activities of full-length FixL to FixJ by ATP-NADH coupled assay (42).
State of heme in FixL [activity]Specific activity
(nmol min−1 mg−1)*
Deoxy (Fe2+) [active]108 ± 4
Oxy (Fe2+-O2) [inactive]43 ± 5
Met (Fe3+) [active]122 ± 6
Cyanomet (Fe3+-CN) [inactive]12 ± 2

*The initial rates were obtained from the time courses for the formation of phosphorylated FixJ. These values were calculated from more than three independent time course experiments.

Oxy FixL was prepared to bind O2 in air to prevent fast autoxidation. The deoxy form accounted for ~30% of the protein in the oxy FixL samples, presumably because of the low affinity of FixL for O2, which was consistent with a previous report (12). Because of the deoxy FixL contaminating the oxy FixL samples, the phosphotransfer activity of the oxy FixL was higher than that of the cyanomet FixL samples, in which all of the proteins were in the inactive state.