Effect of Ectoine, Hydroxyectoine and β-hydroxybutyrate on the Temperature and Pressure Stability of Phospholipid Bilayer Membranes of Different Complexity

Previous research has shown that ectoines fluidize lipid monolayers by increasing the liquid expanded region in DPPC monolayers and also decreasing the line tension responsible for the phase morphology. Here, we explored possible effects of the compatible osmolytes ectoine, hydroxyectoine and β-hydroxybutyrate on lipid bilayer membranes, including effects of temperature and pressure. The effect of the protective osmolytes on the phase transition of DPPC bilayers was investigated by fluorescence spectroscopy, differential scanning calorimetry and pressure perturbation calorimetry. A slight change of the phase behavior was observed, which resulted in a stabilization of the gel phase, which may be caused by an alteration of the hydration properties at the lipid interface and H-bond and electrostatic interactions in the headgroup region. We then explored the cosolvents' effects on giant unilamellar vesicles (GUVs) formed by lipid mixtures exhibiting phase separation into liquid-ordered (l) and liquid-disordered (l) domains using BODIPY-PC and the DiI dye as labels. The presence of both, ectoine and hydroxyectoine showed significant effects on the lateral organization increasing the fluid domains. Moreover, we observed a considerable increase in the adhesion behavior of small vesicles onto GUV surfaces. Diffusion studies by fluorescence recovery after photobleaching experiments on POPC giant vesicles quantitatively showed a hydroxyectoine-induced increase of the diffusion coefficient values, clearly demonstrating an increase in the lateral mobility of lipid within the bilayer membrane. This study provides clear evidence for the fluidizing effect of the compatible solutes on bilayer lipid membranes. A marked effect, however, was only detected if phase separated domains exist.