| 1. |
- Beranová, Lenka, et al.
(author)
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Effect of heavy water on phospholipid membranes : experimental confirmation of molecular dynamics simulations
- 2012
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In: Physical Chemistry, Chemical Physics - PCCP. - : Royal Society of Chemistry. - 1463-9076 .- 1463-9084. ; 14:42, s. 14516-14522
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Journal article (peer-reviewed)abstract
- Although there were experimental indications that phospholipid bilayers hydrated with D(2)O express different biophysical properties compared with hydration by ordinary H(2)O, a molecular concept for this behavior difference was only recently proposed by a molecular dynamics simulations study [T. Róg et al., J. Phys. Chem. B, 2009, 113, 2378-2387]. Here we attempt to verify those theoretical predictions by fluorescence measurements on 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) membranes. Specifically, we determine the water isotope effect on headgroup hydration and mobility, lateral lipid diffusion and lipid backbone packing. Time-dependent fluorescence shift experiments show significantly slower dynamics and lower hydration of the headgroup region for a bilayer hydrated with D(2)O, an observation in good agreement with the calculated predicted differences in duration of lipid-lipid and lipid-water bridges and extent of water penetration into the bilayer, respectively. The water isotope effect on the lipid order parameter of the bilayer core (measured by fluorescence anisotropy) and lateral diffusion of lipid molecules (determined by two-focus fluorescence correlation spectroscopy) is close to the experimental errors of the experiments, however also refers to slightly more rigid organization of phospholipid bilayers in heavy water. This study confirms the view that the water isotope effect can be particularly found in time-resolved physicochemical properties of the membrane. Together with the simulations our experiments provide a comprehensive, molecular view on the effect of D(2)O on phospholipid bilayers.
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| 2. |
- Humpolickova, Jana, et al.
(author)
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Dynamics and size of crosslinking-induced lipid nanodomains in model membranes
- 2012
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In: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 102:3, s. 294a-
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Journal article (peer-reviewed)abstract
- Changes of membrane organization upon crosslinking of its components trigger cellsignaling response to various exogenous factors. Crosslinking of raft gangliosides GM1with cholera toxin (CTxB) was demonstrated to cause microscopic phase separation inmodel membranes and the CTxB-GM1 complexes forming a minimal lipid raft unit aresubject of ongoing cell membrane research. Yet, those subdiffraction sized rafts havenever been described in terms of size and dynamics. By means of two-color z-scanfluorescence correlation spectroscopy, we show that the nano-sized domains are formedin model membranes at lower sphingomyelin content than needed for the large scalephase separation and that the CTxB-GM1 complexes are confined in the domains poorlystabilized with sphingomyelin. Fluorescence resonance energy transfer together withMonte Carlo modeling of the donor decay response reveal the domain radius ofapproximately 8 nm, which increases at higher sphingomyelin content. We observed twotypes of differently behaving domains, which suggests a dual role of the crosslinker: first,local transient condensation of the GM1 molecules compensating lack of sphingomyelinand second, coalescence of existing nanodomains ending in large scale phase separation.
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| 3. |
- Sachl, Radek, 1982-, et al.
(author)
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Fluorescence Study of the Solvation of Fluorescent Probes Prodan and Laurdan in Poly(ε-caprolactone)-block-poly(ethylene oxide) Vesicles in Aqueous Solutions with Tetrahydrofurane
- 2008
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In: Langmuir. - : American Chemical Society (ACS). - 0743-7463 .- 1520-5827. ; 24:1, s. 288-295
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Journal article (peer-reviewed)abstract
- Steady-state and time-resolved fluorescence measurements were used to study the relaxation of the microenvironmentof hydrophobic probes 6-propionyl-2-(dimethylamino)naphthalene (prodan) and 6-dodecanoyl-2-(dimethylamino)-naphthalene (laurdan) in systems containing vesicles formed by the amphiphilic diblock copolymer poly(-caprolactone)-block-poly(ethylene oxide) (PCL-PEO) and water/tetrahydrofurane (THF) solvent mixtures. It was found that in caseof prodan, both steady-state and time-resolved emission spectra were composed of two subspectra corresponding tothe emission of prodan molecules located (i) in fairly rigid (effectively viscous) and hydrophobic domains of thevesicles close to the PCL/PEO interface and (ii) in a more polar and less viscous medium (in the bulk solution). Thefraction of the emission from the more polar microenvironment increases with increasing content of THF in the system.Laurdan, in contrast to prodan, appeared to be solubilized preferentially in the hydrophobic domains up to 30 vol %of THF content, and its emission spectra changed only due to swelling of hydrophobic PCL domains by added THF.The study shows that the analysis of the time-resolved emission from a probe distributed in two media is, in principle,possible, but it is quite complex and appreciably less accurate, and the relaxation times are ill-defined averages ofseveral processes. The bimodal or shoulder-containing time-resolved spectra have to be decomposed in pertinenttime-resolved subspectra and treated separately. Another important result of the study is a piece of knowledge concerningthe motion of the probe with respect to the vesicle. In the studied complex system, not only the relaxation of the solventand reorganization of polymer segments around the fluorescent headgroup of the probe affect the emission but alsoa lateral motion of the probe with respect to the nanoparticle within the lifetime of the excited state contributessignificantly to the relaxation and to the relatively slow time-resolved Stokes shift.
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| 4. |
- Sachl, Radek, 1982-, et al.
(author)
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Limitations of electronic energy transfer in the determination of lipid nanodomain sizes
- 2011
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In: Biophysical Journal. - : Elsevier. - 0006-3495 .- 1542-0086. ; 101:11, s. L60-L62
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Journal article (peer-reviewed)abstract
- Even though superresolution microscopy indicates that size of plasma membrane rafts is <20 nm, those structures have never been observed. Forster resonance energy transfer (FRET) is therefore still the most powerful optical method for characterization of such domains. In this letter we investigate relation between nanodomain affinity of a donor-acceptor (D/A) pair and the detectable nanodomain size/area. We show that probes with high affinity to the liquid-ordered (L(o)) phase are required for detecting domain sizes of a few nanometers, and/or domains that occupy a few percent of the bilayer area. A combination of donors and acceptors that prefer different phases is the more favorable approach. For instance, a D/A pair with the distribution constant of donors K(D) = 5 and acceptors K(A) = 0.01 can resolve a broad spectrum of nanodomain sizes. On the other hand, currently available donors and acceptors that prefer the same phase, either the liquid-disordered (L(d)) or L(o) phase, are not so convenient for determining domain sizes <20 nm. Here the detection limits of FRET experiments employing several commonly used D/A pairs have been investigated.
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| 5. |
- Stefl, Martin, et al.
(author)
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Dynamics and size of cross-linking-induced lipid Nanodomains in model Membranes
- 2012
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In: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 102:9, s. 2104-2113
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Journal article (peer-reviewed)abstract
- Changes of membrane organization upon cross-linking of its components trigger cell signaling response to various exogenous factors. Cross-linking of raft gangliosides GM1 with cholera toxin ( CTxB) was shown to cause microscopic phase separation in model membranes, and the CTxB-GM1 complexes forming a minimal lipid raft unit are the subject of ongoing cell membrane research. Yet, those subdiffraction sized rafts have never been described in terms of size and dynamics. By means of two-color z-scan fluorescence correlation spectroscopy, we show that the nanosized domains are formed in model membranes at lower sphingomyelin (Sph) content than needed for the large-scale phase separation and that the CTxB-GM1 complexes are confined in the domains poorly stabilized with Sph. Forster resonance energy transfer together with Monte Carlo modeling of the donor decay response reveal the domain radius of similar to 8 nm, which increases at higher Sph content. We observed two types of domains behaving differently, which suggests a dual role of the cross-linker: first, local transient condensation of the GM1 molecules compensating for a lack of Sph and second, coalescence of existing nanodomains ending in large-scale phase separation.
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| 6. |
- Valenta, Jan, et al.
(author)
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Light-Emission Performance of Silicon Nanocrystals Deduced from Single Quantum Dot Spectroscopy
- 2008
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In: Advanced Functional Materials. - : Wiley. - 1616-301X .- 1616-3028. ; 18:18, s. 2666-2672
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Journal article (peer-reviewed)abstract
- Spectra of individual silicon nanocrystals within porous Si grains are studied by the wide-field imaging microspectroscopy and their ON-OFF, blinking is detected by the confocal single-photon-counting microscopy. Observed spectral and blinking properties comprise all features reported before in differently prepared single Si nanocrystals (SiNCs). Former apparently contradictory results are shown to be due to different experimental conditions. When the effect of dark periods (OFF switching) is removed the common ultimate photoluminescence properties Of SiO2 passivated SiNCs are found, namely the quantum efficiency (QE) of about 10-20% up to the pumping rate corresponding to one exciton average excitation per quantum dot. At higher pump rates the QE is slowly decreasing as the 0.7th power of excitation. This is most likely due to Auger recombination which, however, seems to be weakened compared with measurements of nanocrystal assemblies. We conclude that SiNCs may be pumped above one exciton occupancy to yield a higher light emission, being advantageous for applications.
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