Unraveling Lipid Membrane Dynamics and Phase Behavior Using AFM

Synthetic lipid bilayers are crucial for modeling cell membranes, as they enable the controlled study of membrane properties and interactions in a simplified, reproducible environment. AFM-based force spectroscopy, in turn, is an ideal technique to investigate the mechanical properties of lipid bilayers at the nanoscale, their elastic mudulus [1], but also their deformation and rupture [2].<br/><br/>In lipid membranes, the ultimate lipid phase coexistence to be fully understood is transient nanodomains, often (confusedly) referred to as lipid rafts [3]. Based on current knowledge, microdomains in equilibrium are no longer considered suitable models for the biological structure that rafts represent. Multiscale spatiotemporal measurements of membrane mechanical properties can help to experimentally address different scenarios where membrane micro- and nanodomain formations provide theoretical support. AFM-based force spectroscopy can resolve the coexistence of domains at concentrations where height differences at domain boundaries are not detectable [4], providing an ideal approach for investigating the mechanical properties of lipid bilayers at the nanoscale. High-speed AFM imaging provides information about the dynamics of domain boundaries. Here, we will discuss several examples of non-equilibrium membrane fluctuations. First, the in situ conversion of sphingomyelin to ceramide. Ceramide is produced in cells from sphingomyelin by means of the enzymatic activity of endogenous sphingomyelinase, impacting the physicochemical properties of the membrane and inducing changes in the curvature, phase, segregation, and order. Then, we will discuss the effect of antimicrobial compounds. Mag2 and PGLa are two antimicrobial peptides that, upon their interaction with biomembranes, have been shown to gradually insert into the lipid bilayer as heterodimer clusters inducing several membrane perturbations, such as alterations in lipid packing, pore openings, and membrane disintegration. Finally, we will address microbial glycolipids, surfactants that can integrate into the microbial cell membranes due to their amphiphilic nature, disrupting the integrity of the membrane. Using these examples, we will conclude that AFM measurements to explore the nanoscale mechanical properties and dynamic behavior of lipid bilayers enhance our understanding of membrane structure and function.<br/><br/>[1] L. Redondo-Morata, R. L. Sanford, O. S. Andersen et al, Biophys J, 111 (2016), p. 363.<br/>[2] L. Redondo-Morata, P. Losada-Pérez, M.I. Giannotti, Curr Top Membr, 86 (2020), p.1.<br/>[3] F. M. Goñi, Chem Phys Lipids, 218 (2019), p. 34.<br/>[4] L. Redondo-Morata et al., Langmuir, 28 (2012), p. 12851.