Browsing by Author "Balcomb, Blake Howard"

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    The Staphylococcus aureus redoxome : characterization of enzymes involved in oxidative stress resistance and survival
    (Stellenbosch : Stellenbosch University, 2019-12) Balcomb, Blake Howard; Erick, Strauss; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: The prevalence of antibiotic resistance is increasing at an alarming rate in South Africa, as well as globally. In particular, multi-drug resistant pathogens such as Staphylococcus aureus (MRSA) are a significant health concern, as these are responsible for the majority of hospital- and community-acquired infections. The human innate immune system is the first line of defense against such pathogens. An important component of these defenses is the chemical attacks lodged by various white blood cells; these attacks include the release of oxidising reagents such as hydrogen peroxide (H2O2) and hypochlorous acid (HOCl), the latter also being the active component of domestic bleach. In order to protect itself against these attacks S. aureus uses a network of redox enzymes and low molecular weight (LMW) thiols, many of which have been shown to play crucial roles in defending against reactive oxygen species. However, the manner in which the effects of HOCl is counteracted is not as well understood. This project set out to characterize enzymes of unknown function that appear to be involved in oxidative stress resistance. In addition, the mechanism by which selected LMW thiols protect a key metabolic enzyme in S. aureus from oxidative damage was also investigated. The results of the study uncover for the first time the activity of a previously uncharacterized enzyme that is conserved in several pathogenic bacteria, and identify it as a key actor in S. aureus’s defence mechanisms against HOCl, as well as the reactive chlorine species that form in the human body when it is released by neutrophils. Furthermore, a small collection of compounds were synthesized and examined as potential mechanism-based inhibitors of this enzyme. Considering the enzyme’s importance as a bacterial defense mechanism at the human-microbe interface, this discovery provides an exciting opportunity for novel drug development focused on the host-pathogen interaction. The results of this study also demonstrate for the first time the moonlighting role of coenzyme A (CoA) — usually a central metabolic cofactor — in protecting enzymes under oxidative stress conditions. In addition, a novel function of S. aureus CoA disulfide reductase was proposed. Finally, the mechanism whereby the LMW thiol bacillithiol is recycled in S. aureus was also explored. Taken together, the results of this study demonstrate the importance of understanding basic molecular mechanisms at the host-pathogen interface that may provide new insight and avenues for novel drug development.