Gram-negative bacteria comprise the majority of microbes that cause infections that are resistant to pre-existing antibiotics. The complex cell wall architecture contributes to their ability to form biofilms, which are often implicated in hospital-acquired infections. Biofilms promote antibiotic resistance by enabling the bacteria to survive hostile environments such as UV radiation, pH shifts, and antibiotics. The outer membrane of Gram-negative bacteria contains lipopolysaccharide (LPS), which plays a role in adhesion to surfaces and formation of biofilms. The main focus of this work was the synthesis of a library of glycolipids designed to be simplified analogues of the Lipid A, the membrane embedded portion component of LPS, to be tested as substrates or inhibitors of Heptosyltransferase I (HepI or WaaC, a glycosyltransferase enzyme involved in the biosynthesis of LPS). Fourteen analogues were synthesized successfully and characterized. While these compounds were designed to function as nucleophilic substrates of HepI, they all demonstrated mild inhibition of HepI. Kinetic characterization of inhibition mechanism identified that the compounds exhibited uncompetitive and mixed inhibition of HepI. Since both uncompetitive and mixed inhibition result in the formation of an Enzyme-Substrate-inhibitor complex, molecular docking studies (using AutoDock Vina) were performed, to identify potential allosteric binding site for these compounds. The inhibitors were shown to bind to a pocket formed after undergoing a conformational change from an open to a closed active site state. Inhibition of HepI via an allosteric site suggest that disruption of protein dynamics might be a viable mechanism for the inhibition of HepI and potentially other enzymes of the GT-B structural class.

in me, for your unwavering support and for your patience those times I couldn't make it to our Skype dates. You are pretty amazing. Anenyasha, thank you for the long phone conversations and 'hangouts', they kept me sane during my thesis writing. To my fellow Zimbabweans at Wes: Matenga, Oscar and Iris, thank you guys for your much needed company. Sometimes you just want to eat sadza, speak Shona and listen to some good old Tuku or Macheso music. Upon request, I would also like to thank my HAS lab buddy Nelson (I refuse to call you King) for keeping me company through most of my thesis writing. Above of all I give thanks the Lord for making it all possible. He has brought me this far... Ebenezer.