Translocation of Phospholipids and Dithionite Permeability in Liquid-Ordered and Liquid-Disordered Membranes

Moreno, Maria João; Estronca, Luís M.B.B.; Vaz, Winchil L.C.

doi:10.1529/biophysj.106.082115

Cited by 48 publications

(57 citation statements)

References 41 publications

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“…61 One minute following quencher addition to the suspensions, a 55.42% (head-labeled) or 47.53% (tail-labeled) decrease in total fluorescence intensity was observed (Figure 8). Dithionite is a charged molecule (dithionite anion) and has very low permeability across lipid bilayers 62 ; therefore, it can only quench the fluorescence of lipid molecules placed in the outer leaflet of the bilayer. No further decrease in fluorescence intensities was observed upon addition of more quencher to the suspensions.…”

Section: Resultsmentioning

confidence: 99%

Lipid Bilayers Covalently Anchored to Carbon Nanotubes

Dayani

Malmstadt

2012

Langmuir

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The unique physical and electrical properties of carbon nanotubes make them an exciting material for applications in various fields such as bioelectronics and biosensing. Due to the poor water solubility of carbon nanotubes, functionalization for such applications has been a challenge. Of particular need are functionalization methods for integrating carbon nanotubes with biomolecules and constructing novel hybrid nanostructures for bionanoelectronic applications. We present a novel method for the fabrication of dispersible, biocompatible carbon nanotube-based materials. Multi-walled carbon nanotubes (MWCNTs) are covalently modified with primary amine-bearing phospholipids in a carbodiimide-activated reaction. These modified carbon nanotubes have good dispersibility in nonpolar solvents. Fourier transform infrared (FTIR) spectroscopy shows peaks attributable to the formation of amide bonds between lipids and the nanotube surface. Simple sonication of lipid-modified nanotubes with other lipid molecules leads to the formation of a uniform lipid bilayer coating the nanotubes. These bilayer-coated nanotubes are highly dispersible and stable in aqueous solution. Confocal fluorescence microscopy shows labeled lipids on the surface of bilayer-modified nanotubes. Transmission electron microscopy (TEM) shows the morphology of dispersed bilayer-coated MWCNTs. Fluorescence quenching of lipid-coated MWCNTs confirms the bilayer configuration of the lipids on the nanotube surface and fluorescence anisotropy measurements show that the bilayer is fluid above the gel-to-liquid transition temperature. The membrane protein α-hemolysin spontaneously inserts into the MWCNT-supported bilayer, confirming the biomimetic membrane structure. These biomimetic nanostructures are a promising platform for the integration of carbon nanotube-based materials with biomolecules.

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Section: Resultsmentioning

confidence: 99%

Lipid Bilayers Covalently Anchored to Carbon Nanotubes

Dayani

Malmstadt

2012

Langmuir

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“…3A). The NBD-SDT reaction rate is much higher than the SDT membrane permeation, NBD-dipalmitoylphosphatidylethanolamine (NBD-DPPE) translocation, and photo bleaching rates (26). When we dissolved the SUVs with detergent, 20 mM Triton X100, we observed another fast drop in the fluorescence to zero due to the exposure of the remaining inner leaflet NBD-labeled lipids to the SDT, which is still abundant in the buffer (Fig.…”

Section: Measurement Of the Nbd-labeled Lipid Transverse Distributionmentioning

confidence: 92%

“…SDT undergoes an irreversible reduction reaction with the NBD and makes NBD nonfluorescent. Furthermore, SDT has very low permeability through the membrane bilayer (25,26). Prior to these measurements, the SUVs were diluted 10-fold in NaCl-HEPES buffer and incubated for 14 to 16 hours at 37 • C to allow the NBD-labeled lipids to equilibrate across the bilayer (confirmed by the quenching measurements).…”

Section: Measurement Of the Nbd-labeled Lipid Transverse Distributionmentioning

confidence: 99%

Measurement of the membrane curvature preference of phospholipids reveals only weak coupling between lipid shape and leaflet curvature

Kamal

Mills

Grzybek

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

126

115

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In biological processes, such as fission, fusion and trafficking, it has been shown that lipids of different shapes are sorted into regions with different membrane curvatures. This lipid sorting has been hypothesized to be due to the coupling between the membrane curvature and the lipid's spontaneous curvature, which is related to the lipid's molecular shape. On the other hand, theoretical predictions and simulations suggest that the curvature preference of lipids, due to shape alone, is weaker than that observed in biological processes. To distinguish between these different views, we have directly measured the curvature preferences of several lipids by using a fluorescence-based method. We prepared small unilamellar vesicles of different sizes with a mixture of egg-PC and a small mole fraction of N-nitrobenzoxadiazole (NBD)-labeled phospholipids or lysophospholipids of different chain lengths and saturation, and measured the NBD equilibrium distribution across the bilayer. We observed that the transverse lipid distributions depended linearly on membrane curvature, allowing us to measure the curvature coupling coefficient. Our measurements are in quantitative agreement with predictions based on earlier measurements of the spontaneous curvatures of the corresponding nonfluorescent lipids using X-ray diffraction. We show that, though some lipids have high spontaneous curvatures, they nevertheless showed weak curvature preferences because of the low values of the lipid molecular areas. The weak curvature preference implies that the asymmetric lipid distributions found in biological membranes are not likely to be driven by the spontaneous curvature of the lipids, nor are lipids discriminating sensors of membrane curvature.curvature coupling | lipid spontaneous curvature | small unilamellar vesicle | fluorescence quenching I n biological membranes, lipids are not distributed homogeneously. The inhomogeneity of lipid distributions within a bilayer may be due to preferential association of particular lipid types with membrane proteins (1, 2), domain formation as a result of the interactions between lipids and sterols (3) or due to the preferences of lipids of different shapes toward regions of matching membrane curvature (4). Membrane regions in which the lipid inhomogeneity or sorting is observed are often highly curved. For example, (i) during mating in the protozoon Tetrahymena thermophila, the pore-containing membrane regions, which become highly negatively curved during fusion, are enriched in coneshaped 2-aminoethylphosphonolipids (5); (ii) in Ca 2+ -dependent exocytosis and in regulated synaptic vesicle fusion, cone-shaped phosphatidylinositol-4,5-bisphosphate lipids are important (6, 7), and are thought to facilitate the formation of an intermediate fusion structure by localizing at the fusion site (8); and (iii) lipids of different shapes are differently sorted in the tubules and vesicles in the endosomal pathway (9). These observations have raised the possibility that the distribution of the lipids may be de...

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“…This electrochemical process also discards the use of chemical reducing agentsthat may undergo slow internalization within vesicles. [53] Furthermore,t he FLIE procedure appeared reliable to explore the reorganization dynamics of as ingle leafleti nr eal time. The capacity to totally bleach the fluorophores located in the outer leaflet also enables the possibility to monitor their passage between inner and outer leaflets (e.g.…”

Section: Resultsmentioning

confidence: 99%

Selective Electrochemical Bleaching of the Outer Leaflet of Fluorescently Labeled Giant Liposomes

Jimenez

Challier

Delacotte

et al. 2017

Chemistry A European J

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Electrochemistry and confocal fluorescence microscopy were successfully combined to selectively bleach and monitor the fluorescence of NBD (7-nitrobenz-2-oxa-1,3-diazole)-labeled phospholipids of giant liposomes. Three types of giant unilamellar vesicles have been investigated, the fluorescent phospholipids being localized either mainly on their outer-, inner-, or both inner/outer leaflets. We established that only the fluorescent lipids incorporated in the outer leaflet of the vesicles underwent electrochemical bleaching upon reduction. The relative fluorescence intensity decay was quantified all along the electrochemical extinction through an original fluorescence loss in electrobleaching (FLIE) assay. As expected, the reorganization of the fluorescent phospholipids followed diffusion-driven dynamics. This was also evidenced by comparison with fluorescence loss in photobleaching (FLIP) and the corresponding numerical model. The value of the lateral diffusion coefficient of phospholipids was found to be similar to that obtained by other methods reported in the literature. This versatile and selective bleaching procedure appears reliable to explore important biological and pharmacological issues.

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Translocation of Phospholipids and Dithionite Permeability in Liquid-Ordered and Liquid-Disordered Membranes

Cited by 48 publications

References 41 publications

Lipid Bilayers Covalently Anchored to Carbon Nanotubes

Lipid Bilayers Covalently Anchored to Carbon Nanotubes

Measurement of the membrane curvature preference of phospholipids reveals only weak coupling between lipid shape and leaflet curvature

Selective Electrochemical Bleaching of the Outer Leaflet of Fluorescently Labeled Giant Liposomes

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