Solid domains in lipid vesicles and scars

The free energy of a crystalline domain coexisting with a liquid phase on a spherical vesicle may be approximated by an elastic or stretching energy and a line tension term. The stretching energy generally grows as the area of the domain, while the line tension term grows with its perimeter. We show that if the crystalline domain contains defect arrays consisting of finite length grain boundaries of dislocations (scars) the stretching energy grows linearly with a characteristic length of the crystalline domain. We show that this result is critical to understand the existence of solid domains in lipid-bilayers in the strongly segregated two phase region even for small relative area coverages. The domains evolve from caps to stripes that become thinner as the line tension is decreased. We also discuss the implications of the results for other experimental systems and for the general problem that consists in finding the ground state of a very large number of particles constrained to move on a fixed geometry and interacting with an isotropic potential.Comment: 7 pages, 6 eps figure

Parameterization of a coarse-grained model of cholesterol with point-dipole electrostatics

© 2018, Springer Nature Switzerland AG. We present a new coarse-grained (CG) model of cholesterol (CHOL) for the electrostatic-based ELBA force field. A distinguishing feature of our CHOL model is that the electrostatics is modeled by an explicit point dipole which interacts through an ideal vacuum permittivity. The CHOL model parameters were optimized in a systematic fashion, reproducing the electrostatic and nonpolar partitioning free energies of CHOL in lipid/water mixtures predicted by full-detailed atomistic molecular dynamics simulations. The CHOL model has been validated by comparison to structural, dynamic and thermodynamic properties with experimental and atomistic simulation reference data. The simulation of binary DPPC/cholesterol mixtures covering the relevant biological content of CHOL in mammalian membranes is shown to correctly predict the main lipid behavior as observed experimentally

Probing lipid mobility of raft-exhibiting model membranes by fluorescence correlation spectroscopy

Confocal fluorescence microscopy and fluorescence correlation spectroscopy (FCS) have been employed to investigate the lipid spatial and dynamic organization in giant unilamellar vesicles (GUVs) prepared from ternary mixtures of dioleoyl-phosphatidylcholine/sphingomyelin/ cholesterol. For a certain range of cholesterol concentration, formation of domains with raft-like properties was observed. Strikingly, the lipophilic probe 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI-C-18) was excluded from sphingomyelin-enriched regions, where the raft marker ganglioside GM1 was localized. Cholesterol was shown to promote lipid segregation in dioleoyl-phosphatidylcholine-enriched, liquid-disordered, and sphingomyelin-enriched, liquid-ordered phases. Most importantly, the lipid mobility in sphingomyelin-enriched regions significantly increased by increasing the cholesterol concentration. These results pinpoint the key role, played by cholesterol in tuning lipid dynamics in membranes. At cholesterol concentrations &gt; 50 mol%, domains vanished and the lipid diffusion slowed down upon further addition of cholesterol. By taking the molecular diffusion coefficients as a fingerprint of membrane phase compositions, FCS is proven to evaluate domain lipid compositions. Moreover, FCS data from ternary and binary mixtures have been used to build a ternary phase diagram, which shows areas of phase coexistence, transition points, and, importantly, how lipid dynamics varies between and within phase regions.</p

SYBP 4.11: Poster

Currently, very few strategies of sample preparation for single molecule analysis exist. Open volume techniques have the disadvantage that molecules can only be detected for a short time, while immobilization techniques, such as tethering the molecules to a surface or embedding them in gels, may perturb the function of molecules. Here we propose a novel technique of capturing single molecules that combines the major advantages of the currently available approaches. Through use of lipid vesicles the size of a confocal volume element, we can confine the movement of molecules to the spatial dimensions of a focal spot without the need for chemical linkers. Thus, the observable kinetics is mainly determined by the photophysics of the chromophore. Used in conjunction with a laser trap, standard measurements, such as TCSPC, can be performed with the added benefit that the microenvironment surrounding the molecules can be controlled and individually varied. This technique can be extended for use in measuring enzyme reactions kinetics where the confined volume may provide advantageous diffusional constraints to promote reactions not favored in dilute solution

Scanning dual-color cross-correlation analysis for dynamic co- localization studies of immobile molecules

Dual-color fluorescence cross-correlation analysis has proven to be a powerful tool to probe interactions of different molecular species in solution and living cells on a single molecule level. Probing the coordinated motion of molecules through the measurement volume, it is a much more selective and data-compressing alternative to co-localization analysis by dual-color imaging, and provides additional access to fast internal dynamics of the co-migrating molecules. However, cellular FCS applications often suffer from extremely low molecular mobility, introducing bleaching artifacts or entirely impeding fluctuation analyses of any kind. Thus, to meet the increasing demand for interaction measurements of nearly stationary molecules, such as receptor-ligand complexes on cell membranes, these limitations of conventional fluorescence correlation and cross-correlation analysis need to be overcome. This can be achieved by combining a piezo-driven stage scanning unit with the confocal FCS setup, minimizing the photodynamic strain imposed on immobile single molecules without compromising the relevant cross-correlation information. Different scanning patterns were chosen and the corresponding auto- and cross-correlation curves recorded for both in vitro and in vivo systems. Expectedly, the shape of the correlation curves depends crucially on the different modes of stage motion. Nevertheless, cross-correlation amplitudes clearly reflect on the presence or absence of linkages between the different molecular species. Marked differences between bound and unbound single molecules could be observed on immobilized proteins in PAA gels and on cell membranes