GLRCE: Research Projects on MRSA
Research Project 3 (GLRP003) : Therapies for anthrax and MRSA.
Bacterial growth is dependent on the acquisition of iron from the surrounding environment. In the Gram-positive pathogens Bacillus anthracis and Staphylococcus aureus, this process relies heavily on the biosynthesis of siderophores. These small secondary metabolites are excellent chelating agents and we apply both genetic and biochemical approaches to discovering how these compounds are synthesized and utilized by the cell to scavenge for iron and promote pathogen survival.
Petrobactin is a "stealth siderophore" produced by B. anthracis which evades host defenses against similar iron chelators; this molecule is essential for pathogenesis and spore outgrowth in the causative microbe of anthrax. Up-regulation of biosynthetic gene clusters under host-like conditions leads to expression of involved "siderophore synthases". These proteins form complexes establishing unique non-ribosomal peptide synthetase (NRPS)-independent machinery reliant on multiple condensation reactions for production of a secondary metabolite. Previous characterization of products encoded by the Bacillus asb operon has demonstrated petrobactin only requires three simple, diffusible precursors: 3,4-dihydroxybenzoic acid, spermidine, and citrate. The petrobactin biosynthetic pathway is however more complex than first anticipated, with multiple routes toward the complete siderophore's synthesis demonstrated in vitro. Additionally, in recent collaboration with GLRCE members at the Joachimiak and Hanna labs, the origin of 3,4-DHBA, the unusual moiety conferring petrobactin's stealth ability and iron-chelating effect has been determined through mechanistic and structural elucidation of the dehydroshikimate dehydratase AsbF (structure pictured).
Lessons learned from petrobactin have been applied to study secondary metabolic reactions by iron-restricted S. aureus strains. Unique activity of individually purified biosynthetic proteins may prove applicable in future engineering of new compounds, creation of useful microbial strains, or isolation of inhibitors for iron sequestration in virulent species. Upon characterization of the pathways pathogens rely on for iron homeostasis within the host, we search for candidate inhibitors of these processes by capitalizing on the expanding chemical libraries housed onsite at the Center for Chemical Genomics within the Life Sciences Institute. Results of these studies indicate further characterization of siderophore biosynthesis and utilization pathways to be promising avenues in exploring new therapeutics capable of shutting down nutrient acquisition mechanisms associated with dangerous bacterial infection.
Research Project 6 (GLRP006) : Therapies against MRSA SuperAntigens
Staphylococcal enterotoxins and toxic shock syndrome toxin-1 (TSST-1) are exceptionally toxic to humans, with doses as low as 0.1 ug/human causing symptoms of toxic shock syndrome through superantigen activities. The major staphylococcal superantigens associated with serious human diseases include TSST-1 and enterotoxin serotypes A-E and Q. This project, through the collaborative interaction of investigators at the University of Minnesota and University of Illinois, develops soluble high-affinity T cell receptor chains that neutralize the toxicities of these superantigens, as measured in rabbit models of diseases. Rabbit models include intravenous, subcutaneous, and intra-pulmonary administration of both superantigens and high-affinity T cell receptor chains. The serum half-life of each high-affinity T cell receptor chain and ability to rescue animals from superantigen-induced serious illness will be assessed. Studies are also planned to evaluate the ability of the high-affinity T cell receptor chains to neutralize superantigens made in vivo in rabbits by methicillin-resistant S. aureus, which have recently been determined to be the most significant causes of serious infectious diseases in the United States.
