Characterization and Target Identification of Hydrazine carbonothiols, a class of trans-Translation Inhibitors

Open Access
Author:
Miller, Sydney Alexis
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Kenneth Keiler, Thesis Supervisor
  • David Scott Gilmour, Honors Advisor
Keywords:
  • trans-translation
  • hydrazine carbonothiols
  • antibiotic resistance
Abstract:
Antibiotic resistance is a growing global epidemic with many common bacteria acquiring mutations to be multi-drug resistant. In order to combat this problem, novel antibiotics have to be developed. One promising target is trans-translation, which is a mechanism used by bacteria to rescue ribosomes stalled during translation. Inhibitors of trans-translation produce a build-up of stalled ribosomes that are unable to translate mRNA, resulting in reduced production of proteins. This is toxic to bacterial cells and leads to cell death. One class of inhibitors identified by the Keiler lab is the hydrazine carbonothiols. These compounds are shown to have broad-spectrum activity at low concetrations and have minimal toxicity in macrophages. These characteristics make them ideal candidates for further drug development. However, the mechanism of action of this class of drugs was still unknown. In order to determine the target, a chemical biology approach was utilized. This method utilizes a probe analogous to the structure of the hydrazine carbonothiols that can be photo-affinity labeled to the target of interest in the cell and can then be identified by click conjugation to a secondary fluorescent probe. Mass spectrometry analysis of the fluorescent band identified the target of these compounds to be ribosomal protein L7/L12. Initial validation studies provide minimal support for this target; thus, such experiments are critically important for future studies. By confirming the target of this class of compounds, the hydrazine carbonothiols can move onto the next stage of antibiotic development in order to curb the number of antibiotic resistant bacteria.