Spin-label probes

Hyde, J. S. and W. K. Subczynski. 1984. Simulation of ESR spectra of the oxygen-sensitive spin-label probe CTPO. J. Magn. Reson. 56 125-130. 

A review of the application of ESR to the study of free radical polymerisation is given by Yamada and co-workers [146]. A survey of the application ESR spectroscopy spin label/probe methods in heterogeneous polymer systems is provided by Veksli and co-workers [147]. Spin probe methods allow the study of the MD of the polymer, its free volume, phase separation and phase morphology. 

Chin, J.H., and D.B. Goldstein. 1997. Membrane disordering action of ethanol Variation with membrane cholesterol content and depth of the spin label probe. Mol Pharmacol 13 435. 

Miziorko HM, Lane MD, Weidman SW. 3-Hydroxy-3-Methyl-glutaryl coenzyme-A synthase-use of a spin-labeled probe to study acyl coenzyme-A binding. Biochemistry 1979 18 399-403. 

Halestrap initially concluded that the increases in mitochondrial pyruvate transport and carboxylation were due to an increase in A pH secondary to stimulation of the electron transport chain in the cytochrome fee, region [255]. The conclusion was based largely on spectral measurements of the redox state of these cytochromes in the control and stimulated states. The spectral measurements were later found to be artifactual due to low amplitude Ca swelling of the mitochondria. Halestrap then suggested that the stable changes in the mitochondria might reside in the lipid components of the membrane due to phospholipase A 2 activity [261,262], but he has been unable to confirm this with lysophospholipid measurements [263]. On the other hand, using an EPR spin label probe of the lipid environment of the isolated mitochondria, Hoek has found differences between control and treated mitochondria [264]. 

Simatos et al. (1981) measured the mobility of a spin-label probe, TEMPO, a stable free radical commonly used for electron paramagnetic resonance (EPR) spectroscopy. She found that the probe showed no mobility below a critical that correlated to Wq. A critical a also existed at which the probe demonstrated a partitioning into a dissolved and a solid-like state. This critical a could represent the moisture content correlating to Tg, though this concept had not been introduced in foods at that time. The partitioning of a... 

Monolayer Studies of Pure Nitroxide Fatty Acid Spin-Label Probes... 

T he use of spin-label probes to investigate cell membrane structure and function clearly demonstrates the fluidity of membrane lipid structures (1,2,3,4) however, a spin-label probe sees only its immediate environment. Predictions (5, 6, 7, 8) and data (9, 10, 11, 12) show that the introduction, for example, of a substituted oxazolidine ring as part of a typical amphiphatic lipid molecule can also significantly perturb a normal lipid environment. Consequently, some quantitative observations that used spin-label techniques need revision while others may be reduced to the level of qualitative predictions. 

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. 

This presentation, based solely on pure film studies, shows that the validity of fluidity and polarity profiles as determined from spin-label probes, is questionable. More important, however, is the fascinating insight into the unique surface chemistry of these compounds. 

A number of ESR experiments have been performed using spin label probes to investigate the effects of cholesterol. It is concluded that above T, a decrease in fluidity of the phospholipid bilayer occurs with increasing cholesterol content [41-43]. 

O Keeffe, D. H., Ebel, R. E., and Peterson, J. A. (1978). Studies of the oxygen binding site of cytochrome P-450. Nitric oxide as a spin-label probe. /. Biol. Chem. 253, 3509-3516. Pous, C., Giroud, J. P., Damais, C., Raichvarg, D., and Chauvelot-Moachon, L. (1990). Effect of recombinant human interleukin-1 and tumor necrosis factor a on liver cytochrome P-450 and serum a-l-acid glycoprotein concentrations in the rat. Drug Metab. Dispos. 18, 467-470. 

A similar result was obtained In a related study of frog muscle labeled with enriched glycine (46) and Chinese hamster ovary cells loaded with phosphoryl [Me- C] choline (50). These values are considerably less than that (>500 cp) obtained by Keith and Snipes (53) for chlamydomonas on the basis of an EPR spin label Study. The validity of the latter result has been questioned by Finch and Harmon (54) who suggested that the correlation time of the spin label probe could be a weighted average of Its predominant occupation of a "mobile" site and a small occupation of a highly immobilized one. 

Regen (1974) described a method of assessing the mobility of the resin sites by covalent attachment of a spin-label probe [2,2,6,6-tetramethyl-4-piperidinyl-l-oxy group (Tempo) (Fig. 3-1. R = (P)—)] to the chlo-romethylated co(polystyrene-DVB). Rotational correlation times (t) were calculated from observed room-temperature electron paramagnetic resonance (epr) spectra. The degree of swelling, q, (swelled volume/ dry volume) was determined from the measured density of the dry resin and the weight of the imbibed solvent. The data indicate that those... 

Pioneering work on surfactantdipid motions in vesicles was carried out some 30 years ago by McConneh and coworkers at Stanford University using esr spin label probes. Komberg... 

Figure 4 EPR spectra of a labelled reactive topical skin protectant with 12- and 7-doxyl in selected controls and exposed samples. The graph shows a plot of EPR signal integral (relative units) obtained from the labelled formulation as a function of H-MG concentrations for spin-label probes, 5-, 7-, 12-, and 16-doxyl stearic acids.

In Section II.A, it was pointed out that the Arrhenius plots of sugar transport rates have discontinuities at the same temperature at which spin-label probes of the bulk lipid phase undergo discontinuous changes. This implies a sensitivity on the part of the protein to changes in the mobility (fluidity) of membrane lipids. In this section, we will examine this point in greater detail. 

One of the most powerful methods for investigation the structure, spatial organization and physical-chemical properties of complex and supramolecular systems on the microscopic, molecular level is EPR spectroscopy in its spin label/probe technique variant (Berliner, 1976 Buchachenko Wasserman, 1976 Likhtenstein, 1976). Usually, nitroxide radicals of different structure were used for studying of structural peculiarities of ionic liquids and the mobility of spin probes in them. We will discuss shortly the most important results obtained by different authors below. 

King ME, Stavens BW, Spector AA (1977) Diet-induced changes in plasma membrane fatty add composition affect physical properties detected with a spin-label probe. Biochemistry 16 5280- 3283... 

Experiments primarily from the laboratory of Aripov on a CTX isolated from Asian cobra venom also show a direct CTX interaction with phospholipid bilayers. Using spin label probes,the CTX (designated cytotoxin was shown to partially insert into phospholipid bilayers to cause an increase in the order parameter of PA liposomes (58). The CTX also interacts with PC/PS liposomes to cause aggregation, increased membrane permeability and enhanced fusion (59). Insertion of CTX into a PC bilayer containing 10% PA was also shown by x-ray small angle scattering (60). 

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