The Design, Synthesis, and Evaluation of a New Class of Antibiotic

Biotin protein ligase (BPL) is a ubiquitous enzyme that catalyzes the conjugation of biotin and ATP to give biotinyl-5’-AMP 1.02, a key intermediate in the activation of biotin dependent enzymes that are crucial to the survival of all cells. Inhibition of this enzyme critically damages bacteria and thus presents an important strategy to develop a new class of antibiotic. Described in this thesis is the design, synthesis, and biological assay of potent inhibitors of Staphylococcus aureus biotin protein ligase (SaBPL) with a 1,2,3-triazole or sulfonamide based isostere. Chapter One highlights the structure and catalytic mechanism of the target enzyme SaBPL. A summary of the chemistry and in vitro biology of known 1,2,3-triazole-based and sulfonamidebased inhibitors, designed to mimic the natural intermediate biotinyl-5’-AMP 1.02, is also discussed. Chapter Two presents work on the development of a new N1-diphenylmethyl-triazole scaffold to replace the adenosine of biotinyl-5’-AMP 1.02. A series of N1-diphenylmethyl-triazole analogues were designed and prepared, with the guidance of in silico docking, to maximise interactions with the active site of SaBPL. This manifested in significant in vitro potency improvements over previous lead triazole-based inhibitors, where 12 and 13, with imidazole and aminopyridine groups respectively, being the most potent inhibitors reported to date for this chemotype (Ki = 6.01 ± 1.01 and 8.43 ± 0.73 nM). Triazole 12 also demonstrated the best antimicrobial activity reported to date for the triazole-based SaBPL inhibitors against S. aureus ATCC 49775, exhibiting a minimum inhibitory concentration (MIC) of 1 μg/mL. Chapter Three presents a series of potent N1-diphenylmethyl-triazole-based analogues targeted to exploit crucial hydrogen bond interactions within the adenine binding site of SaBPL. The inhibitory activity against SaBPL strongly correlated with the in silico docking, particularly analogues that were proposed to hydrogen bond to Asn212 and Ser128 within the adenine binding site. For example, the butanamide substituent of 14 was predicted by docking to hydrogen bond these key amino acids, which manifested in a Ki value of 10.2 ± 2.4 nM. Chapter Four describes the design, detailed characterisation by NMR, and biological assay of a new 1,2,3-triazole-based inhibitor design. Inclusion of a carbonyl at the C10 position of a benzyl-triazole-based SaBPL inhibitor (1.07) gave analogue 4.02, which was shown by docking to hydrogen bond Lys187 within the phosphate binding region. This presumed hydrogen bonding interaction reflected the enhanced Ki value exhibited by 4.02 against SaBPL, relative to triazole 1.07 (Ki = 0.12 ± 0.01 vs 0.25 ± 0.03 μM, respectively). Chapter Five is concerned with the optimisation of the acidity of central NH comprising the sulfonyl linker of the highly effective sulfonamide based SaBPL inhibitors. Acidity of the central NH has been proposed to influence SaBPL inhibitory activity, and based on this, a series of sulfonylurea and sulfonylcarbamate linked analogues were prepared. The relative central sulfonyl NH acidity of these compounds was assessed by 1H NMR, with inhibitor activity against SaBPL shown to correlate with the relative acidity of the central NH. In particular, the acidic sulfonylcarbamate analogue (5.76) exhibited an excellent Ki value of 10.3 ± 3.8 nM, whilst also demonstrating potent whole cell activity against S. aureus ATCC 49775 (MIC = 4 μg/mL). Chapter Six reports details of the biological protocols, docking method, and experimental synthetic procedures for compounds described throughout Chapters, Four, and Five.Thesis (Ph.D.) -- University of Adelaide, School of Physics, Chemistry and Earth Sciences, 202