Hybrid biomimetic platforms based on amphiphilic block copolymers

Cell membranes are of great importance in numerous biological processes, and have been widely studied in the field of biochemistry and biophysics. That is the reason why it is crucial to understand the membrane properties in terms of morphology and architecture, together with the functions of each membrane component. For this purpose, artificial membranes have been developed and amphiphilic block copolymers represent ideal candidates for preparing these membranes. Polymeric membranes provide robustness and stability, and can mimic biological membranes, because they self-assemble in bilayers in aqueous environment. Thus, they have been employed for industrial and biomedical applications, such as catalysis, drug screening, biosensing, etc. In this thesis, we design and create solid-supported membranes by combining different diblock and triblock amphiphilic block copolymers, specifically PDMS-b-PMOXA, by using different membrane preparation methods. Membrane properties were characterized by surface analytical techniques: isotherms of the monolayer at air-water interface were recorded; morphology, homogeneity and thickness of the different membranes were analyzed by AFM and force spectroscopy was employed for the mechanical properties of the polymer membranes; QCM-D was employed for quantifying the amount of biomolecules combined with membranes and the viscoelastic properties of the platform before and after the biomolecule recombination; the functionality of the combined biomolecules was monitored with the fluorimetry. In the first part of the thesis, we investigate the field of hybrid membranes: the combination of copolymers and phospholipids led to a new generation of biomimetic materials that combines the mechanical resistance and chemical tunability of the polymers with the fluidity and biocompatibility of the lipids. Interestingly, when the physical state of polymer and lipids is not miscible, they separate in the membrane and form phase domains, which are similar to lipid raft found in biological membranes and are responsible for key functions in the cell (e.g. signalling, receptor trafficking). For the hybrid membrane preparation we deposited polymer-lipid monolayers onto silica supports by the controllable Langmuir-Blodgett transfer technique. Then we investigated the effect of different combination strategies on the accessibility of the model protein cyt c: either spontaneous insertion or covalent attachment through EDC/sNHS chemistry. Finally, we evaluated the peroxidase-like activity of cyt c after combining it with the membrane. We found that the phase domain separation was crucial for facilitating and controlling the protein recombination into a specific membrane domain (polymeric or lipidic). 24 The attachment strategy revealed to be better in terms of protein accessibility and also enhanced the cyt c activity. In the second part of the thesis we explored the solvent-assisted method. This method was previously employed for preparing lipid membranes and we applied it on amphiphilic block copolymers here for the first time. We optimized the experimental conditions (e.g. polymer characteristics, solvent flow rate) and we combined the obtained membranes with different biomolecules. The combination was performed through the specific biotin-avidin interaction and was used to bind enzymes and DNA strands onto the membrane, preserving their functionality also in this case. In the third part of the thesis, we added a fundamental research which was focused on combining the two main concepts of this thesis: we employed the SA method to prepare hybrid membranes composed of block copolymers and phospholipids in a fast and easy way