Description
The focus of this thesis is to investigate the impact of high curvature events on lipid membrane molecular structure and dynamics with advanced biophysical tools including differential scanning calorimetry (DSC), transmission electron microscopy (TEM), and solid- state nuclear magnetic resonance (SSNMR). One class of peptides that are of interest and shown to induce highly curved membranes are Inhibitor Cystine Knot (ICK) peptides extracted from spider venom. These ICK peptides can modulate various ion channels with high affinity and selectivity while possessing resistance to temperatures, enzymatic degradation, and extreme pH, properties that make these ICK peptides a prime target for biomedical therapeutics. One ICK peptide that is explored is GsMtx-4 extracted from the venom of tarantulas (Grammostola rosea). The goal is to elucidate the lipid-peptide interactions and to understand the organization of lipids while GsMTx-4 is embedded in model lipid bilayers. The results reveal that increasing the GsMTx-4 concentration in bilayers lead to the formation of highly curved regions and sub-structures including cylindrical micelles and nanodiscs. SSNMR shows that although the peptide is embedded in the bilayer, it remains primarily at the membrane surface near the lipid glycerol backbone. To further understand lipid bilayer molecular organization under high curvature events, we explored two methods to prepare highly curved DMPC bilayers: (1) physically stabilize high curvature by adsorbing lipid vesicles to the surface of silica nanoparticles and (2) chemically dope DMPC with DHPC to induce highly curved vesicles including cylindrical micelles and nanodiscs. Adsorption of lipid bilayers to form spherically supported vesicles (SSVs) on silica nanoparticles provides a route to systematically control curvature by varying the nanoparticle diameter. These SSVs exhibit a biphasic phase structure where both lamellar and cubic phases are observed by SSNMR with strong evidence for lipid interdigitation when adsorbed to the smallest nanoparticles (~ 4 nm). Lastly, highly curved cylindrical micelle and discoidal bilayers are explored by combining a long (DMPC) and short (DHPC) acyl chain lipid in varying ratios. Overall, this thesis provides routes to produce and characterize highly curved lipid bilayer structures through ICK peptide introduction, vesicle adsorption to silica nanoparticles and via heterogenous DMPC/DHPC lipid mixtures.
