Lipopolysaccharides (LPS) released from the outer membrane (OM) of Gram negative bacteria form aggregates which trigger an in ammatory response in humans and can result in severe septic shock. LPS molecules are heterogenic, macroamphiphilic oligosaccharides with a pro-in ammatory activity strictly related to the colloidal structures formed in solution; the morphology of the colloids formed by LPS is dependent upon the molecular conformation of the LPS monomers. The intact LPS molecule (smooth chemotype) forms elongated micelles in solution whereas truncated LPS molecules (rough chemotypes) form lamellar aggregates. Biological toxicity tests revealed that the dierent chemotypes have dierent toxic activities on cultured macrophage cells. It is believed that the micellar aggregates are responsible for the toxic eect of LPS, while the lamellar phase should not trigger an in ammatory response. The data presented in this thesis revealed that there might be a more complex connection between the toxicity of LPS and the aggregate structures. The amphiphilic cationic antimicrobial peptides (CAPs) LL37 and LFb possess putative anti-endotoxic activities and were therefore studied using small-angle neutron scattering (SANS) and cryo transmission electron microscopy (cryo-TEM) to assess any modication induced to the colloidal structure of LPS, ideally from a high toxic state to a lower toxic morphology of the aggregates. The peptides LL37 and LFb are able to induce changes to the aggregates from dierent LPS chemotypes in relation to their conformation and the LPS molecular structure. Neutron reectivity experiments on monolayer and bilayer membrane models containing dierent chemotypes of LPS were performed in the presence of the peptides; the data revealed that the two peptides have different mechanisms of interaction with membrane models containing LPS. These investigations provide new insight into the structure-activity of LPS in conditions which are more biologically-relevant than previously published data. Additionally the activity of CAPs on membrane models of the Gram negative bacteria OM was investigated to elucidate the dierent mechanisms of interaction and their possible anti-in ammatory role.
|Date of Award
|Richard Harvey (Supervisor) & James Mason (Supervisor)