Decoding the Mechanism of Action for Triazole and 8-hydroxyquinoline trans-Translation Inhibitors
Open Access
- Author:
- Ryan, Taryn Jeanne
- Area of Honors:
- Biochemistry and Molecular Biology
- Degree:
- Bachelor of Science
- Document Type:
- Thesis
- Thesis Supervisors:
- Kenneth Charles Keiler, Thesis Supervisor
Dr. Teh-hui Kao, Thesis Honors Advisor
Dr. Sarah Ellen Ades, Faculty Reader - Keywords:
- trans-translation
antibiotic resistance
8-hydroxyquinoline
triazole
luciferase-based
click chemistry - Abstract:
- Antibiotic resistant infections pose a serious risk to society. In the United States alone, over 2 million people acquire resistant infections per year, and approximately 23,000 people die from these infections annually. Antibiotic resistance is widespread and the necessity to address the problem is apparent. Novel antibiotics with unexplored mechanisms of action are required to circumvent resistant mechanisms. trans-Translation provides a prospective mechanism for drug development. It is a ribosome- rescue pathway present in bacteria and absent in humans. It is required for the viability or virulence of pathogenic bacteria and provides a target for development of novel antibiotics. The modes of action in bacteria for 1,2,4-substituted triazolyl derivatives and 8-hydroxyquinoline inhibitors of trans-translation remain unclear. In these experiments the molecular target of the triazole compound KKL-1005 was identified in E. coli using chemical biology techniques; it was determined to be the ribosomal protein L7/L12. Furthermore, KKL-1005 was shown to not inhibit translation despite its binding to a ribosomal protein. 8-hydroxy-5-nitroquinoline (HNQ), the quinoline compound characterized in the experiments, was shown to inhibit the protease ClpP in vivo, in vitro, and a binding site was predicted using in silico methods. In vivo inhibition of proteolysis was assessed using a mCherry reporter assay. In vitro inhibition of ClpP specifically was assessed using a fluorogenic peptide. In silico docking studies were performed to predict a likely binding site for HNQ. Lastly, development of a new luciferase-based assay was started. Overall, the data point to promising leads for antibiotic development.