Biomembrane Fabrication by the Solvent-assisted Lipid Bilayer (SALB) Method

In order to mimic cell membranes, the supported lipid bilayer (SLB) is an attractive platform which enables in vitro investigation of membrane-related ...

Nanyang Technological University

Widely used in catalysis and biosensing applications, aluminum oxide has become popular for surface functionalization with biological macromolecules, including lipid bilayer coatings. However, it is difficult to form supported lipid bilayers on aluminum oxide, and current methods require covalent surface modification, which masks the interfacial properties of aluminum oxide, and/or complex fabrication techniques with specific conditions. Herein, we addressed this issue by identifying simple and robust strategies to form fluidic lipid bilayers on aluminum oxide. The fabrication of a single lipid bilayer coating was achieved by two methods, vesicle fusion under acidic conditions and solvent-assisted lipid bilayer (SALB) formation under near-physiological pH conditions. Importantly, quartz crystal microbalance with dissipation (QCM-D) monitoring measurements determined that the hydration layer of a supported lipid bilayer on aluminum oxide is appreciably thicker than that of a bilayer on silicon oxide. Fluorescence recovery after photobleaching (FRAP) analysis indicated that the diffusion coefficient of lateral lipid mobility was up to 3-fold greater on silicon oxide than on aluminum oxide. In spite of this hydrodynamic coupling, the diffusion coefficient on aluminum oxide, but not silicon oxide, was sensitive to the ionic strength condition. Extended-DLVO model calculations estimated the thermodynamics of lipid-substrate interactions on aluminum oxide and silicon oxide, and predict that the range of the repulsive hydration force is greater on aluminum oxide, which in turn leads to an increased equilibrium separation distance. Hence, while a strong hydration force likely contributes to the difficulty of bilayer fabrication on aluminum oxide, it also confers advantages by stabilizing lipid bilayers with thicker hydration layers due to confined interfacial water. Such knowledge provides the basis for improved surface functionalization strategies on aluminum oxide, underscoring the practical importance of surface hydration.

Self-assembly formation of lipid bilayer coatings on bare aluminum oxide: overcoming the force of interfacial water.

With increasing temperature, biological macromolecules and nanometer-sized aggregates typically undergo complex and poorly understood reconfigurations, especially in the adsorbed state. Herein, we demonstrate the strong potential of using localized surface plasmon resonance (LSPR) sensors to address challenging questions related to this topic. By employing an LSPR-based gold nanodisk array platform, we have studied the adsorption of sub-100-nm diameter 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) lipid vesicles on titanium oxide at two temperatures, 23 and 50 °C. Inside this temperature range, DPPC lipid vesicles undergo the gel-to-fluid phase transition accompanied by membrane area expansion, while DOPC lipid vesicles remain in the fluid-phase state. To interpret the corresponding measurement results, we have derived general equations describing the effect of deformation of adsorbed vesicles on the LSPR signal. At the two temperatures, the shape of adsorbed DPPC lipid vesicles on titanium oxide remains nearly equivalent, while DOPC lipid vesicles become less deformed at higher temperature. Adsorption and rupture of DPPC lipid vesicles on silicon oxide were also studied for comparison. In contrast to the results obtained on titanium oxide, adsorbed vesicles on silicon oxide become more deformed at higher temperature. Collectively, the findings demonstrate that increasing temperature may ultimately promote, hinder, or have negligible effect on the deformation of adsorbed vesicles. The physics behind these observations is discussed, and helps to clarify the interplay of various, often hidden, factors involved in adsorption of biological macromolecules at interfaces.

Contribution of temperature to deformation of adsorbed vesicles studied by nanoplasmonic biosensing.

Complement activation plays an important role in innate immune defense by triggering formation of the membrane attack complex (MAC), which is a biomacromolecular assembly that exhibits membrane-lytic activity against foreign invaders including various pathogens and biomaterials. Understanding the details of MAC structure and function has been the subject of extensive work involving bulk liposome and erythrocyte assays. However, it is difficult to characterize the mechanism of action of MAC inhibitor drug candidates using the conventional assays. To address this issue, we employ a biomimetic supported lipid bilayer platform to investigate how two MAC inhibitors, vitronectin and clusterin, interfere with MAC assembly in a sequential addition format, as monitored by the quartz crystal microbalance-dissipation (QCM-D) technique. Two experimental strategies based on modular assembly were selected, precincubation of inhibitor and C5b-7 complex before addition to the lipid bilayer or initial addition of inhibitor followed by the C5b-7 complex. The findings indicate that vitronectin inhibits membrane association of C5b-7 via a direct interaction with C5b-7 and via competitive membrane association onto the supported lipid bilayer. On the other hand, clusterin directly interacts with C5b-7 such that C5b-7 is still able to bind to the lipid bilayer, and clusterin affects the subsequent binding of other complement proteins involved in the MAC assembly. Taken together, the findings in this study outline a biomimetic approach based on supported lipid bilayers to explore the interactions between complement proteins and inhibitors, thereby offering insight into MAC assembly and regulation.

Saziye Yorulmaz

School of Materials Science and Engineering,
Centre for Biomimetic Sensor Science

Nanyang Technological University

Biomembrane Fabrication by the Solvent-assisted Lipid Bilayer (SALB) Method

Saziye Yorulmaz

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Nanyang Technological University

Biomembrane Fabrication by the Solvent-assisted Lipid Bilayer (SALB) Method

School of Materials Science and Engineering,
Centre for Biomimetic Sensor Science