Infectious disease is a global threat to human health, thus defining an urgent need to develop novel antimicrobial strategies that limit the impact of life-threatening bacterial infections. The parent project supports our fundamental studies of bacterial transition metal (manganese, copper and zinc) homeostasis, sulfur metabolism and sulfide homeostasis to accelerate the pace of discovery of new antibacterial targets. In this proposal, we seek to obtain instrumentation that will allow us to probe how metal trafficking proteins, or metallochaperones, allow a bacterium to adapt to ?remodeling? of transition metal bioavailability under culture conditions that mimic host-mediated metal starvation, or ?nutritional immunity.? We propose to leverage our emerging expertise in the proteomics of the Staphylococcus aureus and Enterococcus faecalis developed under the parent project to elucidate how currently uncharacterized, COG0523 family GTPase metallochaperones impact the metalloproteome in these organisms. Partial funding is requested to acquire an integrated Perkin-Elmer NexION 2000C inductively coupled plasma-mass spectrometry (ICP-MS)-Flexar liquid chromatography (LC) system to establish a state-of-the-art metalloproteomics workflow in my laboratory. This instrumentation will be used to replace an existing ICP-MS that is no longer serviceable, and which lacks the LC-ICP-MS integration, high sensitivity, extended dynamic range and simultaneous multi-element ionization schemes characteristic of the new instrumentation, and required by this workflow. Use of this instrumentation will yield new insights into the cellular mechanisms of adaptation to the vertebrate host defense against invading microorganisms, fundamental to transition metal biology and future therapeutics discovery efforts.
In this proposal, we request supplemental funding to acquire an integrated Perkin-Elmer NexION 2000C ICP-MS Flexar LC system to establish a state-of-the-art metalloproteomics (speciation) workflow in my laboratory. This instrumentation will be used to investigate the dynamics of transition metal (re)- distribution among cellular proteins under conditions of host-mediated ?nutritional immunity? against important bacterial pathogens.
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