Date of Award

5-2023

Document Type

Thesis

Department

Biochemistry

Committee Chair/Advisor

Dr. James Morris

Abstract

Kinetoplastid parasite infections remain a global health burden. Here, we have characterized inhibitors of an essential Trypanosoma brucei glycolytic enzyme, enolase (TbENO). We anticipate TbENO inhibitors will be potent anti-trypanosomals, as T. brucei relies on glycolysis for ATP production in the blood of infected mammals. Additionally, the phosphonate enolase inhibitors being considered are well-tolerated in mammals, suggesting their potential in developing novel therapeutics for kinetoplastid infections. TbENO was cloned into the bacterial expression vector, pQE-30, and the heterologously-expressed protein was purified by nickel affinity and assayed in a coupled enzyme assay. Enzyme activity paralleled the abundance of a ~45 kDa protein, consistent with the predicted size of TbENO. Preliminary inhibition assays have demonstrated that phosphonate-based enolase inhibitors could be promising leads for blocking the glycolytic pathway in the pathogen. For example, (1-hydroxy-2-oxopyrrolidin-3-yl) phosphonic acid (deoxy-SF2312) was a potent enzyme inhibitor (IC50 value of 0.60 ± 0.23 µM), while larger ring-bearing variants were less potent. Molecular docking simulations supported this supposition, as larger ring-bearing variants were predicted to have lower binding affinities. The inhibitory potency and anti-trypanosomal activity of these compounds suggest they are promising therapeutic leads for future development.


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