Doctoral Degrees (Biochemistry)
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Browsing Doctoral Degrees (Biochemistry) by Subject "Antibiotics"
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- ItemSynthesis and evaluation of inhibitors targeting Coenzyme : a biosynthesis and metabolism in Staphylococcus aureus(Stellenbosch : University of Stellenbosch, 2010-12) Van der Westhuyzen, Renier; Strauss, Erick; University of Stellenbosch. Faculty of Science. Department of Chemistry and Polymer ScienceENGLISH ABSTRACT: The human pathogen Staphylococcus aureus is a major cause of hospital-, and more recently, community-acquired infections. The rate at which this organism is acquiring resistance to antibiotics is increasing while the development of new antibiotics is slowing down. There is therefore a desperate need for new antistaphylococcal agents, and in particular ones with novel mechanisms of action that can be used to circumvent established resistance pathways. Unlike humans, S. aureus employs the essential cofactor coenzyme A (CoA) as its major low molecular weight thiol. Together, CoA and the enzyme CoA disulfide reductase (CoADR) are responsible for maintaining the internal redox homeostasis in this organism, and disruption of this balance (or reduction of CoA levels) may therefore be potential mechanisms by which new antistaphylococcal agents may act. In this study we set out to achieve this by direct inhibition of CoADR, and by inhibition of one or more of the CoA biosynthetic enzymes. For the inhibition of CoADR CoA analogues containing Michael acceptors were designed and prepared by employing a chemo-enzymatic approach. This strategy involved the chemical synthesis of pantothenamides containing α,β-unsaturated ester, ketone and sulfone moieties as Michael acceptors, followed by their biotransformation into the corresponding CoA analogues by three CoA biosynthetic enzymes. The compounds prepared in this manner all inhibited CoADR potently. A full kinetic evaluation of the inhibition by these compounds suggested that these compounds act by alkylation of the single active site cysteine of CoADR in an irreversible fashion. In this study we also set out to determine the mechanism of action of the antistaphylococcal compound CJ-15,801, which is structurally similar to pantothenic acid, the biosynthetic precursor of CoA. Due to this similarity we proposed that the antibiotic properties of CJ-15,801 are based on the inhibition of enzymes involved in CoA biosynthesis and metabolism. Our investigations confirmed that the second enzyme of the CoA pathway, phosphopantothenoylcysteine synthetase (PPCS), acts as the main target of CJ-15,801. These studies were followed by an investigation into alternative synthetic methodologies for the preparation of CJ-15,801 and its analogues. As a result an established Pd-catalyzed coupling reaction was modified and applied in the third known total synthesis of CJ-15,801, as well as of several of its analogues. This protocol has several advantages over its predecessors, most importantly its suitability for preparing these compounds on large (up to one gram) scale.