Stearic acid spin label

Kusumi, A., W. K. Subczynski, and J. S. Hyde. 1982a. Effects of pH on ESR spectra of stearic acid spin labels in membranes Probing the membrane surface. Fed. Proc. 41 1394. 

Penetration of the biomembrane is undoubtedly essential for most membrane activity. Araki and Rifkind (13) obtained esr spectra of stearic acid spin labelled erythrocyte membranes in the presence of diverse compounds including Triton XlOO, chlorpromazine and glutaraldehyde. The two surfactants chlorpromazine and Triton XlOO both increase the rate of haemolysis and are shown to increase membrane fluidity. Glutaraldehyde as expected decreases fluidity and decreases the rate of haemolysis. 

Figure 7 Plot of the change in the product of the coupling and maximum saturation factors as a function of macromolecular structure. At lower pH values, the spin-labelled lipids are present as vesicles and vesicular aggregates, while at higher pH values, micelles are formed. The higher psmax values for the micelles imply greater water accessibility to the radical site. The solid circles represent 16-DS (16-doxyl stearic acid, spin-labelled at the end of the lipid tail) while the open circles represent 5-DS (5-doxyl stearic acid, spin-labelled near the polar head group). Reproduced with permission from Ref. [70].

Godici, P. E., and Landsberger, F. R. (1975). Biochemistry 14, 3927. 1 3C Nuclear Magnetic Resonance Study of the Dynamic Structure of Lecithin-Cholesterol Membranes and the Position of Stearic Acid Spin-Labels. 

Fig. 8.23 ESR spectra of the 14-stearic acid spin label (SASL) in complexes of the K26 peptide with DMPC at Upid/peptide ratios of 4.6,...

Effects of anesthetics on the mobility of stearic acid spin labels in phospholipid vesicles and in mitochondrial membranes... 

Figure 1 Chemical structure of the stearic acid spin labels.

The chromaffin granule from the adrenal medulla has a membrane with high lipid/protein ratio, in line with its function as an exo-cytotic storage vesicle. The ESR spectra of various positional isomers of the stearic acid spin labels (cf. 1(3,12) = 14-SASL, Fig. 3.1), at probe concentrations in these membranes, are given in Fig. 3.2. The total spectral anisotropy of these labels, given by the linesplitting 2(A - ), is seen to decrease as the spin... 

Membranes with a relatively high protein content frequently display a second component in the ESR spectra of lipid spin labels, in addition to the fluid lipid bilayer component discussed in the previous section. This component is best resolved with labels close to the end of the chain, since a large degree of averaged spectral anisotropy is available to detect any immobilization induced by the protein. The ESR spectra of the 16-SASL stearic acid spin label in acetylcholine receptor-rich membranes and in bilayers of the extracted lipids is given in Fig. 3.3. A motionally restricted spin label component is seen in the outer wings of the spectrum from the membranes which is not present in the spectrum from the lipids alone. 

Fig. 3.2. ESR spectra of various positional isomers of the stearic acid spin label l(m,n) [ = (n + 2) - SASL] (ca. 1 wt%) in chromaffin granule membranes at 30 C (Marsh and Watts, 1982 Fretten et al., 1980 Marsh et al., 1976).

Fig. 3.3. ESR spectra of the stearic acid spin label, 16-SASL, in acetylcholine receptor-rich membranes from Torpedo mar-morata. a) Membranes, b) Aqueous dispersion of extracted membrane lipids, c) Immobilized component difference spectrum obtained by subtracting the lipid spectrum (55% of the total integrated intensity) from the membrane spectrum, d) Fluid component difference spectrum obtained by subtracting a purely immobilized spectrum (45% relative intensity) from the membrane spectrum (Marsh and Barrantes, 1978 Marsh et al., 1981).

Fig. 3.5. ESR spectra of the cardiolipin spin label 14-CLSL (top row), stearic acid spin label 14-SASL (middle row) and phosphatidylcholine spin label 14-PCSL (bottom row), in ...

Observed collisions between 14N 15N spin-label pairs are indicated. DMPC and POPC molecules are also shown. POPC represents the major component (70%) of the EYPC mixture, (b) Bimolecular collision rate for a nitroxide moiety at the C16 position of the stearic acid alkyl chain with other SASLs in the DMPC alone and the DMPC with 10mol% lutein at 27°C. (From Yin, J.J. and Subczynski, W.K., Biophys../., 71, 832,1996. With permission.)... 

Oxidation of 2,2,4,4-tetraalkyloxazolidines gives stable nitroxide radicals (265) which are used as spin labels for probing biomolecular structures (69ACR17, B-76MI41800). The stearic acid derivative 12-doxylstearic acid (266) is commercially available. 

Ohtsuru et al. (25) have recently investigated the behavior of phosphatidylcholine in a model system that simulated soy milk. They used spin-labelled phosphatidylcholine (PC ) synthesized from egg lysolecithin and 12-nitroxide stearic acid anhydride. The ESR spectrum of a mixture of PC (250 yg) and native soy protein (20 mg) homogenized in water by sonication resembled that observed for PC alone before sonication. However, when PC (250 yg) was sonicated in the presence of heat-denatured soy protein (20 mg), splitting of the ESR signal occurred. On this basis, they postulated the existence of two phases PC making up a fluid lamella phase and PC immobilized probably due to the hydrophobic interaction with the denatured protein. In a study of a soy-milk model, Ohtsuru et al. (25) reported that a ternary protein-oil-PC complex occurred when the three materials were subjected to sonication under the proper condition. Based on data from the ESR study, a schematic model has been proposed for the reversible formation-deformation of the ternary complex in soy milk (Figure 2). 

ESR spectroscopy has also been used to study dynamics in more complex systems on oxide surfaces, i.e. self-assembled molecular (SAM) films [136, 137]. Fig. 28 shows ESR spectra taken for spin labeled stearic acid, namely n-doxyl-stearic acid (n-DXSA) immersed onto a thin alumina film. These nitrox-ides, with an oxazolidinyl ring as paramagnetic group connected to different positions of the aliphatic chain, are well known as paramagnetic probes in the study of natural and synthetic membranes. The spin labeled molecules are pres-... 

The method of biosynthetic incorporation of spin label, rather than mechanical addition to isolated material, is a convenient way of ensuring that the results obtained are biologically meaningful and has also been used with such systems as the mould Neurospora crassa [158], Mycoplasma laidlawii [159], human leucocytes, and mouse L cells [160]. The spectra from these two mammalian cells showed distinct similarities for a variety of spin labels, but different spectra were obtained when the labels were incorporated in human erythrocytes. Fractionation of the cell components showed the stearic acid (C, n = 3) spin label in all the major fractions, but by far the largest concentration was in the nuclear membrane. The ESR spectrum underwent a time and temperature dependent decay and spin labels on the surface membrane were reactivated with K3Fe(CN)6. 

To understand the behavior of the mixed host-probe films, it is essential to understand the behavior of the pure films of both components. We previously investigated in detail the behavior of the pure films of 12-nitrox-ide stearate [2-( 10-carboxydecyl)-2-hexyl-4,4 -dimethyl-3-oxazolidinyloxyl] including its extraordinary temperature dependence (7, 8, 16). In this paper we extend our investigations to the pure films of other nitroxide stearic acid (and methyl ester) probes where the oxazolidine ring is attached to various carbon atoms of the stearic acid (or ester) hydrocarbon chain. This series of spin-label probes is one of those most extensively used to study cell membrane structure. It was used to define, among other things, an order parameter establishing the fluidity (17) and polarity profiles (18) of lipid bilayers. 

Spin labels are stable, paramagnetic molecules that, by their structme, easily attach themselves to various biological macromolecular systems such as proteins or cell membranes. Examples of spin labels that can be covalently bonded to specific sites of biological systems include nitroxide derivatives of A-ethylmaleimide, which bind specifically to -SH groups, and nitroxide derivatives of iodoacetamide, which bind specifically to methionine, lysine, and arginine residues of amino acids. Nonco-valently bonded spin-labels that can be incorporated into biological systems include nitroxide derivatives of stearic acid, of phospholipids, and of cholesterol. 

In recent years the systems most often studied by the spin-label method have been biological membranes and various models thereof. For example, using nitroxide derivatives of stearic acid, the organization of the phospholipid phase of biological membranes has been studied [6]. One interesting practical application of this type of spin-labeling is in the study of disease-state membranes. Intact erythrocyte mem-... 

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