Effects of hydrogen sulfide on the heme coordination structure and catalytic activity of the globin-coupled oxygen sensor AfGcHK

Af GcHK is a globin-coupled histidine kinase that is one component of a two-component signal transduction system. The catalytic activity of this heme-based oxygen sensor is due to its C-terminal kinase domain and is strongly stimulated by the binding of O 2 or CO to the heme Fe(II) complex in the N-terminal oxygen sensing domain. Hydrogen sulfide (H 2 S) is an important gaseous signaling molecule and can serve as a heme axial ligand, but its interactions with heme-based oxygen sensors have not been studied as extensively as those of O 2 , CO, and NO. To address this knowledge gap, we investigated the effects of H 2 S binding on the heme coordination structure and catalytic activity of wild-type Af GcHK and mutants in which residues at the putative O 2 -binding site (Tyr45) or the heme distal side (Leu68) were substituted. Adding Na 2 S to the initial OH-bound 6-coordinate Fe(III) low-spin complexes transformed them into SH-bound 6-coordinate Fe(III) low-spin complexes. The Leu68 mutants also formed a small proportion of verdoheme under these conditions. Conversely, when the heme-based oxygen sensor Ec DOS was treated with Na 2 S, the initially formed Fe(III)–SH heme complex was quickly converted into Fe(II) and Fe(II)–O 2 complexes. Interestingly, the autophosphorylation activity of the heme Fe(III)–SH complex was not significantly different from the maximal enzyme activity of Af GcHK (containing the heme Fe(III)–OH complex), whereas in the case of Ec DOS the changes in coordination caused by Na 2 S treatment led to remarkable increases in catalytic activity.