Direct Measurement of the Mechanical Properties of Lipid Phases in Supported Bilayers

Abstract : Biological membranes define not only the cell boundaries but any compartment within the cell. To some extent, the functionality of membranes is related to the elastic properties of the lipid bilayer and the mechanical and hydrophobic matching with functional membrane proteins. Supported lipid bilayers (SLBs) are valid biomimetic systems for the study of membrane biophys-ical properties. Here, we acquired high-resolution topographic and quantitative mechanics data of phase-separated SLBs using a recent atomic force microscopy (AFM) imaging mode based on force measurements. This technique allows us to quantitatively map at high resolution the mechanical differences of lipid phases at different loading forces. We have applied this approach to evaluate the contribution of the underlying hard support in the determination of the elastic properties of SLBs and to determine the adequate indentation range for obtaining reliable elastic moduli values. At ~200 pN, elastic forces dominated the force-indentation response and the sample deformation was <20% of the bilayer thickness, at which the contribution of the support was found to be negligible. The obtained Young's modulus (E) of 19.3 MPa and 28.1 MPa allowed us to estimate the area stretch modulus (k A) as 106 pN/nm and 199 pN/nm and the bending stiffness (k c) as 18 k B T and 57 k B T for the liquid and gel phases, respectively. The concept that lipid bilayers are not just a simple passive beholder of membrane proteins is now well accepted. It is important to note that membranes are heterogeneous, with local associations of lipids (and proteins) in detergent-resistant membrane (DRM) domains or rafts (1,2). In general, membrane dimensions and mechanical properties (i.e., bilayer thickness, bending and stretching stiffness, or membrane tension) modify the function not only of mecha-nosensitive proteins but of any membrane protein (3). In this framework, the mattress model is in favor of the importance of the lipid environment and provides an elastic model of lipid bilayer behavior (4). As a consequence of the established importance of bilayer compliance and lateral organization of membranes, a large number of techniques (including micropi-pette aspiration, surface force apparatus, biomembrane force probe or atomic force microscopy imaging, and force spectroscopy) have been employed to give insights into the structure and mechanical properties of biological membranes (5–8). Among them, atomic force microscopy (AFM) (9) in particular has been used to address fundamental questions on the nanomechanics of supported membranes (10). Here, we introduce a novel, to our knowledge, AFM-based imaging technique, PeakForce-Quantitative Nano-Mechanics (PF-QNM), to probe the structural and mechanical properties of SLBs. PF-QNM allows simultaneous imaging and quantitative mechanical mapping of the sample, both at submolecu-lar resolution (11), and, it is important to point out, improves acquisition time and spatial resolution compared to other AFM-based techniques, such as force volume. This is achieved by oscillating the sample in the z axis at a given amplitude (tens of nanometers) and frequency (2 kHz), thus providing cycles of force-distance (FnD) curves in which the tip intermittently contacts the sample surface. Each FnD plot is thereafter analyzed to determine the mechanical properties of the sample (Fig. S1 in the Supporting Material), thus coupling topography analysis with stiffness and deformation assessment at high resolution. The aim of this study was to probe the mechanical properties of biological membranes in the elastic regime. We present measures of the elastic properties (i.e., Young's modulus) of different lipid phases, and characterize the effect of the underlying hard substrate. Nanomechanical mapping of SLBs was performed on DOPC/DPPC (1:1, mol/mol) membranes (Fig. 1), which is one of the best-characterized SLBs and is commonly used as a straightforward model membrane for AFM studies (12,13). DOPC/DPPC bilayers display phase separation at room temperature between liquid (L a) and gel (L b) phase, as a consequence of the different transition temperatures of DOPC and DPPC (À20 C and 41 C, respectively) (14). The presence
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Biophysical Journal, Biophysical Society, 2012, 102 ((1)), pp.L01-3. 〈10.1016/j.bpj.2011.11.4001〉
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Laura Picas, Felix Rico, Simon Scheuring. Direct Measurement of the Mechanical Properties of Lipid Phases in Supported Bilayers. Biophysical Journal, Biophysical Society, 2012, 102 ((1)), pp.L01-3. 〈10.1016/j.bpj.2011.11.4001〉. 〈inserm-01363280〉

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