Mycobacterium tuberculosis DosS binds H(2)S through its Fe(3+) heme iron to regulate the DosR dormancy regulon

Mycobacterium tuberculosis (Mtb) senses and responds to host-derived gasotransmitters NO and CO via heme-containing sensor kinases DosS and DosT and the response regulator DosR. Hydrogen sulfide (H(2)S) is an important signaling molecule in mammals, but its role in Mtb physiology is unclear. We have previously shown that exogenous H(2)S can modulate expression of genes in the Dos dormancy regulon via an unknown mechanism(s). Here, we test the hypothesis that Mtb senses and responds to H(2)S via the DosS/T/R system. Using UV–Vis and EPR spectroscopy, we show that H(2)S binds directly to the ferric (Fe(3+)) heme of DosS (K(D)(app) = 5.30 μM) but not the ferrous (Fe(2+)) form. No interaction with DosT(Fe(2+)-O(2)) was detected. We found that the binding of sulfide can slowly reduce the DosS heme iron to the ferrous form. Steered Molecular Dynamics simulations show that H(2)S, and not the charged HS(−) species, can enter the DosS heme pocket. We also show that H(2)S increases DosS autokinase activity and subsequent phosphorylation of DosR, and H(2)S-mediated increases in Dos regulon gene expression is lost in Mtb lacking DosS. Finally, we demonstrate that physiological levels of H(2)S in macrophages can induce DosR regulon genes via DosS. Overall, these data reveal a novel mechanism whereby Mtb senses and responds to a third host gasotransmitter, H(2)S, via DosS(Fe(3+)). These findings highlight the remarkable plasticity of DosS and establish a new paradigm for how bacteria can sense multiple gasotransmitters through a single heme sensor kinase.