Stability and applications of model membranes

Author: Alex Ashenden

Ashenden, Alex, 2022 Stability and applications of model membranes, Flinders University, College of Science and Engineering

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Whilst biological membranes act as a barrier between the inside and outside cell environment ubiquitously across all organisms, there is a high degree of architectural complexity inherent in the structure. This makes systematic studies in areas such as protein functionality and antibiotic efficacy difficult to perform. Various model systems have been developed to mimic biological membranes using only essential components.

This thesis focuses on one specific type of model system, tethered-bilayer lipid membranes. In a tethered-bilayer lipid membrane a solid supporting structure is used as a base to which anchoring tether lipids bind to form a monolayer, with additional lipids added to complete the bilayer. Spacer lipids are also utilised to bind to the substrate at different ratios to increase bilayer fluidity and sub-membrane hydration. As a planar surface-based model a variety of techniques can be used to analyse these systems such as electrochemical impedance spectroscopy and neutron scattering, which will be the techniques showcased through this thesis.

This thesis is split into three distinct parts. Chapter 1 acts as an introduction to biological membrane systems and the various models that have been utilised to mimic their structure and capabilities, as well as discussing the various pros and cons of each system in order to explain why tBLMs have been utilised for the work showcased in this thesis. This is followed by chapter 2, a discussion of the methods utilised through the thesis.

The second part of the thesis, chapter 3, focuses on improving the understanding of how the functionality of tBLMs is affected by removal from the aqueous solution they are usually situated in. This is important as a case study for determining the feasibility of these model systems in future biosensor devices where they may be subject to such dehydration, as well as to determine their capacity for transport. Hence, studies were undertaken to determine the quality of tBLMs before and after being dried out and rehydrated at different levels of tethering density. Additionally, the improvement of this rehydration process was investigated using polyelectrolytes to coat the bilayers before dehydration. Finally, complexity of the bilayers was increased through addition of cholesterol, ubiquitous across mammalian cell membranes, to the outer leaflet of the bilayer before the dehydration process to determine if behaviour was similar with a more realistic membrane model.

The final part of this thesis, chapter 4, focuses on the biomedical applications of tBLMs designed to replicate bacterial membranes. Firstly, the use of tBLMs mimicking gram-negative bacteria as a screening tool to improve the efficacy of colistin, a last line antibiotic, is discussed. Colistin’s damaging of the bacterial membrane was shown to be improved when functionalised gold nanoparticles were introduced to the bilayer initially, before addition of colistin afterwards. Secondly, the formation of novel tBLMs from bacterial lipid extracts is discussed. Lipids were extracted directly from A. baumannii cultures and used to form tBLMs with and without the inclusion of DHA, to help determine that membrane potential loss is not a likely antimicrobial mechanism of DHA.

Keywords: biophysics, model membrane, tblm, antibiotics, microbial

Subject: Science, Technology and Enterprise thesis

Thesis type: Doctor of Philosophy
Completed: 2022
School: College of Science and Engineering
Supervisor: Ingo Koeper