Synthesis and membrane interactions of spin-label bifunctional reagents

Gaffney, Betty J.; Willingham, Gary L.; Schepp, Robert S.

doi:10.1021/bi00273a027

Cited by 35 publications

(23 citation statements)

References 49 publications

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“…Pjura et al (1984) used 4.3 x 10-6m and Conrad & Singer (1981) used a 6 x 1O-5 M solution. It should be noted that these concentrations are all far below the critical micelle concentration of 4 x 10-3 M. A better agreement of our data is found with values given by Gaffney et al (1983). They measured partition coefficients of spin-labelled fatty acids by e.s.r.…”

Section: Discussionsupporting

confidence: 89%

Membrane solubility of chlorpromazine. Hygroscopic desorption and centrifugation methods yield comparable results

Luxnat¹,

Müller²,

Galla³

1984

Biochemical Journal

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Binding of the positively charged drug chlorpromazine to artificial and erythrocyte bilayer membranes was investigated by the filtration method called hygroscopic desorption [Conrad & Singer (1979) Proc. Natl. Acad. Sci. U.S.A. 76, 5202-5206] and by the conventional centrifugation method. Only minor differences in the partition coefficients were observed using the two methods. Our finding is not consistent with the observation of Conrad & Singer that amphipaths are completely excluded from biological membranes. However, the partition coefficient is dependent on membrane composition, which means dependent on the physical properties of a membrane.

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Section: Discussionsupporting

confidence: 89%

Membrane solubility of chlorpromazine. Hygroscopic desorption and centrifugation methods yield comparable results

Luxnat¹,

Müller²,

Galla³

1984

Biochemical Journal

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“…DISCUSSION Sheetz & Singer (1974) originally proposed that cationic penetrant amphipathic drugs preferentially distributed into the cytofacial monolayer, whereas anionic penetrant and ionic impenetrant amphipathic drugs preferentially distributed into the exofacial monolayer of the erythrocyte plasma membrane. Regardless of the mechanism of selective fluidization of one or the other monolayer, with one exception (Conrad & Singer, 1979 most evidence is consistent with selectivity for individual monolayers by charged amphipaths (Sheetz & Singer, 1974;Bondy & Remien, 1981;Gaffney et al, 1983;Dipple et al, 1982;Houslay et al, 1977Houslay et al, ,1980bHouslay et al, , 1981. Herein, the testing of the hypothesis was extended to LM-cell plasma membranes, which have a transbilayer gradient ofcholesterol and fluidity opposite to that found in erythrocytes and other normal (i.e.…”

Section: Individual Monolayer Structure Of Lm Plasma Membranesmentioning

confidence: 54%

Charged anaesthetics alter LM-fibroblast plasma-membrane enzymes by selective fluidization of inner or outer membrane leaflets

Sweet¹,

Schroeder²

1986

Biochemical Journal

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The functional consequences of the differences in lipid composition and structure between the two leaflets of the plasma membrane were investigated. Fluorescence of 1,6-diphenylhexa-1,3,5-triene(DPH), quenching, and differential polarized phase fluorimetry demonstrated selective fluidization by local anaesthetics of individual leaflets in isolated LM-cell plasma membranes. As measured by decreased limiting anisotropy of DPH fluorescence, cationic (prilocaine) and anionic (phenobarbital and pentobarbital) amphipaths preferentially fluidized the cytofacial and exofacial leaflets respectively. Unlike prilocaine, procaine, also a cation, fluidized both leaflets of these membranes equally. Pentobarbital stimulated 5'-nucleotidase between 0.1 and 5 mM and inhibited at higher concentrations, whereas phenobarbital only inhibited, at higher concentrations. Cationic drugs were ineffective. Two maxima of (Na+ + K+)-ATPase activation were obtained with both anionic drugs. Only one activation maximum was obtained with both cationic drugs. The maximum in activity below 1 mM for all four drugs clustered about a single limiting anisotropy value in the cytofacial leaflet, whereas there was no correlation between activity and limiting anisotropy in the exofacial leaflets. Therefore, although phenobarbital and pentobarbital below 1 mM fluidized the exofacial leaflet more than the cytofacial leaflet, the smaller fluidization in the cytofacial leaflet was functionally significant for (Na+ + K+)-ATPase. Mg2+-ATPase was stimulated at 1 mM-phenobarbital, unaffected by pentobarbital and slightly stimulated by both cationic drugs at concentrations fluidizing both leaflets. Thus the activity of (Na+ + K+)-ATPase was highly sensitive to selective fluidization of the leaflet containing its active site, whereas the other enzymes examined were little affected by fluidization of either leaflet.

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“…Their results might reflect the extensive disorder of the phospholipidic matrix and of the probe itself above the phase‐transition temperature. In our case, the formation of an ordered bilayer structure due to the transmembrane “immobilization”3, 12, 13 of the probe and to the concomitant use of cholesterol has led to an excellent selectivity for the photolabeling on the transmembrane domain of GPA in vesicles.…”

mentioning

confidence: 83%

Mid-Membrane Photolabeling of the Transmembrane Domain of Glycophorin A in Phospholipid Vesicles

et al. 2001

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The topography of membrane-bound proteins at atomic resolution is known only in rare cases. [1] Although the primary amino-acid sequence of glycophorin A (GPA), the major sialoglycoprotein of the human erythrocytes, has been known for more than twenty years and was the first membrane protein sequence elucidated, [2] a three-dimensional picture of the protein is still missing. We have previously developed the photoactivable membrane probe 1. This is a phospholipid with two distal, polar heads (a bola-amphiphile) and carries a photosensitive group (benzophenone) in the middle of a transmembrane chain. It is easily incorporated into DMPC (1,2-dimyristoyl-sn-glycero-3-phosphocoline) vesicles, where it spans the bilayer at least in the presence of physiological concentrations of cholesterol. We demonstrated recently that the tandem use of the probe 1 a and cholesterol (for its ordering effect) in photolabeling experiments on DMPC vesicles led to a remarkable regioselective functionalization and we are presently conducting such studies. With this information, we expect that the present strategy using Au/ Ag(upd) electrodes for halide detection will provide the notable abilities to measure the concentrations of multiple halides simultaneously and to provide specific signals for confirming their identification. Experimental SectionMaterials: Au (99.99 %) shot and Cr-coated tungsten filaments were obtained from Americana Precious Metals (East Rutherford, NJ) and R. D. Mathis (Long, Beach, CA), respectively. Sulfuric acid (double distilled, 98 %) and silver sulfate (c(Cl À ) 0.02 %) were obtained from Aldrich and used as received. KCl, KBr, and KI were obtained from Mallinckrodt and used as received. Au(111) samples were prepared by sequential evaporation of Cr (2 ± 3 nm) and Au (150 nm) onto glass slides and a post-evaporation annealing in a hydrogen flame. [12] Electrochemical measurements: Cyclic voltammetry was conducted with a computer-controlled PAR Model 263A potentiostat. A solution of 0.6 mm Ag 2 SO 4 and 0.1m H 2 SO 4 in deionized water (Millipore, 18.2 MW) was used to deposit the Ag upd adlayer on gold. Modified electrodes were prepared by cycling once in this solution and being removed at a 300 mV versus Ag / Ag 0 under potentiostatic control. [13] The Au(111)/Ag(upd) electrodes were rinsed with deionized water and immersed into a halide test solution for varying amounts of time. The samples were rinsed with water and characterized electrochemically in a 0.6 mm Ag 2 SO 4 /0.1m H 2 SO 4 (aq) solution. A standard three-electrode configuration was used to obtain all CVs, and all potentials are quoted relative to silver wire (Ag /Ag 0 ).

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Synthesis and membrane interactions of spin-label bifunctional reagents

Cited by 35 publications

References 49 publications

Membrane solubility of chlorpromazine. Hygroscopic desorption and centrifugation methods yield comparable results

Membrane solubility of chlorpromazine. Hygroscopic desorption and centrifugation methods yield comparable results

Charged anaesthetics alter LM-fibroblast plasma-membrane enzymes by selective fluidization of inner or outer membrane leaflets

Mid-Membrane Photolabeling of the Transmembrane Domain of Glycophorin A in Phospholipid Vesicles

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