Examples of recent projects
Spincoating of supported membranes
During the past 5 years we have developed a novel procedure to fabricate supported membranes based on hydration of spincoated lipid films. Thin films made by spincoating membrane lipids on solid substrates will organize in stacked lamellar bilayer structures. This dry film has turned out to be a highly useful precursor structure for forming supported bilayers in water. Upon hydration of the dry film in fluid water one or several stacked supported membranes are easily preserved. This preparation methods is very robust and versatile and allows supported membranes to be formed with an arbitrary lipid composition. We have succesfully applied this method to generate membranes with traditionally difficult compositions such as high anionic lipid content. (Langmuir 2004, 20, 9720-9728)
Domain Patterns in Ternary Model Membranes
A number of morphological and statistical aspects of domain formation in singly and doubly supported ternary membranes have been investigated. Such ternary membranes produce macroscopic phase separation in two fluid phases and are widely used as raft models. We find that membrane interactions with the support surface can have a critical influence on the domain shapes if measures are not taken to screen these interactions. Combined AFM and fluorescence microscopy demonstrate small (500 nm) irregular domains and incomplete formation of much larger (5 ím) round domains. These kinetically trapped structures are the result of interactions between the membrane and the support surface, and they can be effectively removed by employing doubly supported membranes under physiological salt concentrations. These decoupled supported membranes display macroscopic round domains that are easily perturbed by fluid shear flow. The system allows a quantitative characterization of domain coarsening upon being cooled into the coexistence region. We determine the domain growth exponent alpha=0.31, which is in close agrement with the theoretical value of 1/3. Analysis of the spatial domain pattern in terms of Voronoi polygons demonstrates a close similarity to equilibrated cellular structures with a maximized configurational entropy (Langmuir 2007, 23, 8135-8141).
Activation of Phospholipase A(2) by Ternary Model Membranes
Formation of liquid-ordered domains in model membranes can be linked to raft formation in cellular membranes. The lipid stoichiometry has a governing influence on domain formation and consequently, biochemical hydrolysis of specific lipids has the potential to remodel domain features. Activation of phospholipase A2 (PLA2) by ternary model membranes with three components (DOPC/DPPC/Cholesterol) can potentially change the domain structure by preferential hydrolysis of the phospholipids. Using fluorescence microscopy, this work investigates the changes in domain features that occur upon PLA2 activation by such ternary membranes. Double-supported membranes are used, which have minimal interactions with the solid support. For membranes prepared in the coexistence region, PLA2 induces a decrease of the liquid-disordered (Ld) phase and an increase of the liquid-ordered (Lo) phase. A striking observation is that activation by a uniform membrane in the Ld phase leads to nucleation and growth of Lo-like domains. This phenomenon relies on the initial presence of cholesterol and no PLA2 activation is observed by membranes purely in the Lo phase. The observations can be rationalized by mapping partially hydrolyzed islands onto trajectories in the phase diagram. It is proposed that DPPC is protected from hydrolysis through interactions with cholesterol, and possibly the formation of condensed complexes. This leads to specific trajectories which can account for the observed trends. The results demonstrate that PLA2 activation by ternary membrane islands may change the global lipid composition and remodel domain features while preserving the overall membrane integrity (Biophysical Journal 94 (2008) 3966–3975).
DNA-Controlled Assembly of Soft Nanoparticles
Immobilization of DNA (encoding) on solid nanoparticles requires surface chemistry, which is well established for gold surfaces but often tedious and not generally applicable for many other inorganic surface materials. While substantial effort has been devoted to expanding surface chemistry techniques for solid nanoparticles, considerably less attention has been given to the development of noncovalent attachment of DNA to soft nanoparticles, like liposomes. Here we report a DNA-controlled assembly of liposomes in solution and on solid supported membranes, this process displays remarkably sharp thermal transitions from an assembled to a disassembled state, allowing application of DNA-controlled liposome assembly for the detection of polynucleotides (e.g., DNA) with single mismatch discrimination power. The method is based on a single DNA strand (contains two lipid membrane anchors), which is able to noncovalently attach to a liposome surface. This design enables detection of biological polynucleotide targets as the complementary strand can be unmodified DNA and RNA strands (J. AM. CHEM. SOC. 2008, 130, 10462–10463)
Nanobubble induced forces
An atomic force microscope and the colloidal probe technique are used to probe the interaction between a hydrophobic particle and a hydrophobic surface in water. The characteristics of the observed force curves strongly suggest that a gas bubble is formed when the particle is moved toward the surface and that the bubble ruptures when the particle subsequently is retracted from the surface. We demonstrate that this type of interaction is not unique for hydrophobic surfaces in water since the interaction between hydrophilic surfaces in air provides very similar force curves. However, the interaction between hydrophobic surfaces vanish if water is replaced by an organic solvent with low polarity. The bridging bubble model is employed to explain the hysteresis observed between approach and retraction force traces and experimental conditions where the hysteresis can be almost eliminated are identified. Finally, it is demonstrated that the hydrophobic interaction is strongly temperature dependent and this dependence can be attributed mainly to the decreasing solubility of air in water with increasing temperature. (ACS Nano; (Article); 2008; 2(9); 1817-1824. DOI: 10.1021/nn800218s)
Single Molecule Protein Ligand Studies
In order to investigate the dynamic strength of the interaction between lung surfactant protein D (SP-D) and different sugars, maltose, mannose, glucose, and galactose, we have used an atomic force microscope to monitor the interaction on a single molecule scale. The experiment is performed by measuring the rupture force when the SP-D-sugar bond is subjected to a continuously increasing force. Under these dynamic conditions, SP-D binds strongest to D-mannose and weakest to maltose and D-galactose. These results differ from equilibrium measurements wherein SP-D exhibits preference for maltose. On the basis of this finding, we propose that the binding of the disaccharide maltose to SP-D, which is energetically stronger than the binding of any of the monosacchrides, alters the structure of the binding site in a way that lowers the dynamic strength of the bond. We conclude that determining the strength of a proteinligand bond under dynamic stress using an atomic force microscope is possibly more relevant for mimicking the actual nonequilibrium physiological situation in the lungs. (Biochemistry 2007, 46, 12231-12237)
