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O2-specific regulation of the ferrous heme-based sensor kinase FixL from Sinorhizobium meliloti and its aberrant inactivation in the ferric form

2003; Elsevier BV; Volume: 304; Issue: 1 Linguagem: Inglês

10.1016/s0006-291x(03)00556-4

1090-2104

Satoru Akimoto, Atsunari Tanaka, Kayako Nakamura, Yoshitsugu Shiro, Hiro Nakamura,

Microbial Fuel Cells and Bioremediation

FixL, a rhizobial heme-based O2-sensing histidine kinase, catalyzes autophosphorylation in the deoxy form at low O2 tension, while the kinase activity is inhibited in the case of the O2-bound form. The present study unambiguously shows that the binding of CO and NO does not significantly inhibit the kinase activity of dithiothreitol (DTT)-reduced ferrous FixL from Sinorhizobium meliloti, which is inconsistent with the spin state mechanism previously reported. Kinase inactivation is caused by aberrant disulfide (S–S) bond formation at Cys301 in the ferric homodimer, which explains these contradictory observations. The addition of DTT cleaved the S–S bond, leading to restoration of kinase activity in the ferric form as well as heme reduction, but, sodium hydrosulfite treatment produced the kinase-inactive deoxy form without S–S cleavage. On the basis of these experimental results, it can be concluded that ferrous FixL discriminates O2 from CO and NO, and signals the O2-bound state by downregulating the phosphoryl transfer reaction.