Date of Award

2023

Degree Name

Pharmaceutical Sciences

College

School of Pharmacy

Type of Degree

M.S.

Document Type

Thesis

First Advisor

Dr. Timothy E Long, Committee Chairperson

Second Advisor

Dr. Hasan Koc

Third Advisor

Dr. Michael Hambuchen

Abstract

Disulfiram, known as Antabuse®, is an oral drug for the treatment of alcohol dependence. Previous studies have indicated that disulfiram (DSF) exhibits antibacterial effects, particularly against Gram-positive bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA). Our study delves into the antibacterial mechanism of DSF in MRSA through High-Pressure Liquid Chromatography (HPLC) metabolomics, investigating the underlying mechanism of DSF effects on thiamine and amino acid metabolism. Thiamine pyrophosphate (TPP) plays a crucial role as a cofactor for critical enzymes such as transketolase, pyruvate dehydrogenase, and 2-oxoglutarate dehydrogenase. These enzymes are integral to the carbohydrate metabolism process within bacterial cells. TPP also contributes to coenzyme A (CoA) biosynthesis, identified as a prospective drug target for DSF in MRSA. Recent research highlighted DSF's role in lowering intracellular CoA levels in MRSA, and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways helped to uncover enriched genes related to the biosynthesis of TPP. Our transcriptome data further illuminated different amino acid metabolism shifts within DSF-treated MRSA. In-depth HPLC investigations utilized various methods to gauge TPP and amino acid levels in DSF-treated MRSA. These HPLC results effectively validated our hypothesis, confirming DSF's influence on increasing cellular levels of TPP and amino acids like glutamate, glutamine, arginine, glycine, β-alanine, and lysine. Notably, our study also revealed diminished cellular levels of aspartate, valine, phenylalanine, and threonine. Our comprehensive study offers further insight on why DSF treatment alters TPP and select amino acid levels. These findings add to our understanding of DSF's antibacterial mechanism in MRSA.

Subject(s)

Disulfiram.

Staphylococcus aureus infections.

Methicillin resistance.

Antibacterial agents.

High performance liquid chromatography.

Thiamin pyrophosphate.

Enzymes.

Coenzymes.

Biosynthesis.

Amino acids -- Metabolism.

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