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Sugars as Prodrugs: Exploiting Sugar Metabolism in Bacteria

Boulanger, Erin Farrell
http://orcid.org/0000-0003-3550-528X
http://rave.ohiolink.edu/etdc/view?acc_num=osu168141685250898

Abstract Details

2023, Doctor of Philosophy, Ohio State University, Biomedical Sciences.
Antibiotic resistance is an urgent public health problem and is associated each year with over a million deaths worldwide. Strategies to limit antibiotic exposures as well as improved practices for infection prevention and containment have been successful in reducing the emergence and spread of antibiotic resistance. Despite these efforts, novel therapeutic strategies are needed. In this dissertation, we investigate the induction of sugar-phosphate toxicities as a novel therapeutic modality to selectively target microbial pathogens. We use Salmonella enterica serovar Typhimurium, as our model pathogen, due to its clinical relevance, tractable genetics, and well-developed mouse models for studying infection. It is also one of the most characterized microbes and many aspects of its physiology and pathogenesis are relevant to other pathogens of the Enterobacteriaceae family for which antibiotic resistance is of paramount concern [i.e., the Carbapenem-resistant Enterobacteriaceae (CRE) and extended spectrum beta-lactamase (ESBL)-producing pathogens]. Here, we have assembled and characterized a collection of Salmonella mutants predicted to suffer sugar-phosphate toxicity due to the absence of targeted enzymes within a variety of sugar-utilization pathways. Elimination of these enzymes coupled with the provision of the appropriate sugar leads to the accumulation of toxic sugar-phosphate intermediates resulting in growth inhibition. These mutations serve as a proxy for small molecule inhibitors of those enzymes that would be used in real-world therapeutic applications. Of the seven mutants tested in vitro, five (galE, galT, rhaD, mtlD, and araD) mutants showed growth inhibition in addition to a fraB mutant reported in earlier work. All but the galT mutant were also attenuated in a mouse model of Salmonella-mediated gastroenteritis. While homologs of galE are widespread among bacteria and in humans, the araD, mtlD, rhaD, and fraB genes are rare in most phyla of bacteria and are not present in humans, thus rendering the encoded enzymes promising narrow-spectrum therapeutic targets. Following the successful identification of genes that cause sugar-phosphate toxicity when mutated, we designed and optimized a high-throughput screening (HTS) protocol to identify small molecule inhibitors of one of our sugar-phosphate targets, FraB. Of the 224,009 compounds screened at Harvard’s ICCB-Longwood Facility, we identified three lead compounds whose activities against wild-type Salmonella, but not fra-deficient Salmonella, are reproducible and have been confirmed biochemically to inhibit the FraB enzyme. This work sets the precedent for additional HTS to identify inhibitors of other sugar-phosphate toxicity targets. In the concluding study of this dissertation, we began to generate hypotheses regarding the molecular mechanism(s) by which sugar-phosphates inhibit growth. We utilized transcriptomics to characterize the gene expression profiles of cells undergoing four different sugar-phosphate toxicities at two different timepoints. STRING and iModulon analyses of our differentially expressed genes supported our understanding for the biological context of these changes. Overall, the work completed in this dissertation establishes the foundation for exploiting sugar-phosphate toxicity as a novel therapeutic modality to employ against Salmonella and other high priority pathogens.
Brian Ahmer, Ph.D. (Advisor)
Venkat Gopalan, Ph.D. (Committee Member)
Vanessa Hale, DVM, MAT, Ph.D. (Committee Member)
John Gunn, Ph.D (Committee Member)
266 p.
Biomedical Research; Microbiology; Molecular Biology
Salmonella, gastroenteritis, colitis, narrow-spectrum antimicrobial, sugar- phosphate, sugar metabolism

Recommended Citations

Citations

  • Boulanger, E. F. (2023). Sugars as Prodrugs: Exploiting Sugar Metabolism in Bacteria [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu168141685250898

    APA Style (7th edition)

  • Boulanger, Erin. Sugars as Prodrugs: Exploiting Sugar Metabolism in Bacteria. 2023. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu168141685250898.

    MLA Style (8th edition)

  • Boulanger, Erin. "Sugars as Prodrugs: Exploiting Sugar Metabolism in Bacteria." Doctoral dissertation, Ohio State University, 2023. http://rave.ohiolink.edu/etdc/view?acc_num=osu168141685250898

    Chicago Manual of Style (17th edition)

osu168141685250898
126
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