Lipid Membrane Probes

The dyes used for probing lipid membranes consist of a fluorophore with a long lipophilic tail. The lipophilic tail inserts itself into the membrane thus locating the fluorophore label on the membrane. These products are used as lipid labels and in cell tracking as part of biophysical studies. There are two main classes of fluorophore, aminostyryls and indocarbocyanines. The most widely used indocarbocya-nine is the 18-carbon derivative of Cy3 known as dil, (3.72). 

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Siu H, Duhamel J, SasaM D, Pincus JL (2010) Nanodomain formation in lipid membranes probed by time-resolved fluorescence. Langmuir 26 10985—10994... 

SALDiTT T, LI c, SPAAR A. Structure of antimicrobial peptides and lipid membranes probed by interface-sensitive X-ray scattering. Biochim Biophys Acta 2006 Sep 1758(9) 1483-98. 

Talley C E and Dunn R C 1999 Single molecules as probes of lipid membrane microenvironments J. Chem. Phys. B 103 10 214-20... 

Hollars C W and Dunn R C 2000 Probing single molecule orientations in model lipid membranes with near-field scanning optical microscopy J. Phys. Chem 112 7822-30... 

Figure 13.3 (a-c) Trajectories of the diffusion motion of a gold nanoparticle probe on a planar lipid membrane, (d) Mean-square displacement plots forthe diffusion shown in (a-c). Adapted from Ref [31] with permission. 

Baumgard, T, Hunt, G., Farkas, E. R., Webb, W. W. and Feigenson, G. W. (2007) Fluorescence probe partitioning between Lo/Ld phases in lipid membranes. Biochim. Biophys. Acta,... 

Subczynski, W. K., A. Wisniewska, J. S. Hyde, and A. Kusumi. 2007b. Tree-dimensional dynamic structure of the liquid-ordered domain in lipid membranes as examined by pulse-EPR oxygen probing. Biophys. J. 92 1573-1584. 

Demchenko AP, Mely Y, Duportail G, Klymchenko AS (2009) Monitoring biophysical properties of lipid membranes by environment-sensitive fluorescent probes. Biophys J 96 3461-3470... 

It is possible, however, that the electrochromic response of some styrylpyridi-nium probes, for example, RH421 (see Fig. 2), is enhanced by a reorientation of the dye molecule as a whole within the membrane. There is a steep gradient in polarity on going from the aqueous environment across the lipid headgroup region and into the hydrocarbon interior of a lipid membrane. Therefore, any small reorientation of a probe within the membrane is likely to lead to a change in its local polarity and hence a solvatochromic shift of its fluorescence excitation spectrum. Such a... 

The previous result is an important one. It indicates that there can be yet another fruitful route to describe lipid bilayers. The idea is to consider the conformational properties of a probe molecule, and then replace all the other molecules by an external potential field (see Figure 11). This external potential may be called the mean-field or self-consistent potential, as it represents the mean behaviour of all molecules self-consistently. There are mean-field theories in many branches of science, for example (quantum) physics, physical chemistry, etc. Very often mean-field theories simplify the system to such an extent that structural as well as thermodynamic properties can be found analytically. This means that there is no need to use a computer. However, the lipid membrane problem is so complicated that the help of the computer is still needed. The method has been refined over the years to a detailed and complex framework, whose results correspond closely with those of MD simulations. The computer time needed for these calculations is however an order of 105 times less (this estimate is certainly too small when SCF calculations are compared with massive MD simulations in which up to 1000 lipids are considered). Indeed, the calculations can be done on a desktop PC with typical... 

B. Hudson and S. A. Cavalier, in Studies ofMembrane Dynamics and Lipid-Protein Interactions with Parinaric Acid in Speclroscopic Membrane Probes Vol. 1 (L. Loew, ed.), CRC Press, Inc., Boca Raton, Florida (1988). 

MD simulations of model membrane systems have provided a unique view of lipid interactions at a molecular level of resolution [21], Due to the inherent fluidity and heterogeneity in lipid membranes, computer simulation is an attractive tool. MD simulations allow us to obtain structural, dynamic, and energetic information about model lipid membranes. Comparing calculated structural properties from our simulations to experimental values, such as areas and volumes per lipid, and electron density profiles, allows validation of our models. With molecular resolution, we are able to probe lipid-lipid interactions at a level difficult to achieve experimentally. 

The modulation of synaptosomal plasma membranes (SPMs) by adriamycin and the resultant effects on the activity of membrane-bound enzymes have been reported [58]. Again DPH was used as fluorescence probe. Adriamycin increased the lipid fluidity of the membrane labeled with DPH, as indicated by the steady-state fluorescence anisotropy. The lipid-phase separation of the membrane at 23.3 °C was perturbed by adriamycin so that the transition temperature was reduced to 16.2 °C. At the same time it was found that the Na+,K+-stimulated ATPase activity exhibits a break point at 22.8 °C in control SPMs. This was reduced to 15.8 °C in adriamydn-treated SPMs. It was proposed that adriamycin achieves this effect through asymmetric perturbation of the lipid membrane structure and that this change in the membrane fluidity may be an early key event in adriamycin-induced neurotoxicity. 

Another example is the perturbing effect of eight calcium channel blockers on membranes prepared from two different lipids [68]. The authors used total lipids from rat brain and synaptosomal membranes. The spin probe was l-palmitoyl-2-stearoyl-phosphatidyl-choline labeled at the doxyl group at the carbon-16 position (16-PC). The apparent order parameter, S, is calculated from the apparent outer (Amax) and inner (Amin) splittings which were directly taken from the ESR spectra. It is used to describe the relative efficiency of the dmgs in perturbing the lipid membrane. 

Abdiche YN, Myszka DG (2004) Probing the mechanism of drug/lipid membrane interactions using biacore. Anal Biochem 328 233-243... 

Membranes exhibit a common stmcture, with lipid molecules arranged in the form of one or more bilayers, or lamellae. Since lipids are generally nonfluo-rescent, lipid-bound fluorophores are an excellent tool to study this environment. These membrane probes are poorly soluble in water, and hence they partition readily into the hydrophobic regions of the membranes. The derivatives of anthroyloxy fatty acids (AF), with the fluorophore 9-anthroic acid esterfied to the 2, 6,9, 12, or 16 position along a fatty acid acyl chain (stearic acid or palmitic acid), are frequently used. The stmcture of an AF probe is shown schematically in Fig. 1... 

These same AF probes have been widely used to study the bilayer stmcture in a variety of lipid membranes [8-12]. Since the fluorophore is attached at a known position along a fatty acid acyl chain, AF probes have been used to evaluate bilayer stmctures as a function of depth. When partitioned into a membrane, the fluorophore is localized deeper in the bilayer as its attachment site to the fatty acid is moved further away from the carboxyl... 

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