| dc.description.abstract | Staphylococcus aureus is a Gram-positive pathogen that contributes to significant morbidity and mortality worldwide. In order to successfully colonize the host, S. aureus requires heme as a nutrient iron source and as a cofactor for multiple cellular processes. Although required for pathogenesis, excess heme is toxic. S. aureus employs a two-component system, the heme sensor system (HssRS), to sense and protect against heme toxicity. Upon activation, HssRS induces the expression of the heme-regulated transporter (HrtAB) to alleviate heme toxicity. The ability to sense and respond to heme is critical for pathogenesis, yet the mechanism of heme sensing remains unknown. Small molecules ‘882 and ‘3981 were previously identified in a screen for activators of HssRS. Through the utilization of Phrt-driven suicide strain, numerous residues were identified within HssRS and Phrt that are required for heme- and ‘882-induced activation. Furthermore, utilization of the suicide strain identified ‘882 as small molecule activator of coproporphyrinogen oxidase (HemY) from Gram-positive bacteria, an enzyme specific to the Gram-positive heme biosynthesis pathway. Activation of HemY induces accumulation of coproporphyrin III and leads to photosensitization of multiple Gram-positive pathogens. In combination with light, HemY activation reduces bacterial burden and tissue ulceration in murine models of skin and soft tissue infection. Thus, small molecule activation of HemY represents an effective strategy for the development of light-based antimicrobial therapies. The other compound of interest, ‘3981, activates HssRS independently of heme accumulation. ‘3981 is toxic to S. aureus; however, derivatives of ‘3981 were synthesized that lack toxicity while retaining the ability to activate HssRS activation. Using ‘3981 derivatives as probes of the heme stress response, bacterial nitric oxide synthase (bNOS) was identified as crucial for the S. aureus heme stress response, providing evidence that nitric oxide synthesis and heme sensing are intertwined. The chemical genetic approach applied herein provides insight into requirements of bacterial signaling, methods for small molecule target identification, and therapeutic strategies exploiting heme biosynthesis for the treatment infectious diseases. | |
