Conversion of a heme-based oxygen sensor to a heme oxygenase by hydrogen sulfide: effects of mutations in the heme distal side of a heme-based oxygen sensor phosphodiesterase (Ec DOS)

摘要

The heme-based oxygen-sensor phosphodiesterase from Escherichia coli (Ec DOS), is composed of an N-terminal heme-bound oxygen sensing domain and a C-terminal catalytic domain. Oxygen (O_2) binding to the heme Fe(II) complex in Ec DOS substantially enhances catalysis. Addition of hydrogen sulfide (H_2S) to the heme Fe(III) complex in Ec DOS also remarkably stimulates catalysis in part due to the heme Fe(III)-SH and heme Fe(II)-O_2 complexes formed by H_2S. In this study, we examined the roles of the heme distal amino acids, M95 (the axial ligand of the heme Fe(II) complex) and R97 (the O_2 binding site in the heme Fe(II)-O_2 complex) of the isolated hemebinding domain of Ec DOS (Ec DOS-PAS) in the binding of H_2S under aerobic conditions. Interestingly, R97A and R97I mutant proteins formed an oxygenincorporated modified heme, verdoheme, following addition of H_2S combined with H_2O_2 generated by the reactions. Time-dependent mass spectroscopic data corroborated the findings. In contrast, H_2S did not interact with the heme Fe(III) complex of M95H and R97E mutants. Thus, M95 and/or R97 on the heme distal side in Ec DOS-PAS significantly contribute to the interaction of H_2S with the Fe(III) heme complex and also to the modification of the heme Fe(III) complex with reactive oxygen species. Importantly, mutations of the O_2 binding site of the heme protein converted its function from oxygen sensor to that of a heme oxygenase. This study establishes the novel role of H_2S in modifying the heme iron complex to form verdoheme with the aid of reactive oxygen species.