Electromechanical stability of planar lipid membranes from bipolar lipids of the thermoacidophilic archebacterium Sulfolobus acidocaldarius.

Stable planar membranes have been obtained from the bipolar lipid glycerol dialkyl nonitol tetraether (GDNT) extracted from the thermoacidophilic archebacterium Sulfolobus acidocaldarius. The electric capacity Cm, the resistance Rm and tension sigma of these membranes were measured. The dependence of the bipolar lipid membranes mean life time tau 1 on voltage was investigated. It was shown that the irreversible electric breakdown of membranes from GDNT and usual phospholipids is due to the same mechanism, viz., due to formation of a hydrophilic pore with an overcritical radius. Under electric field the GDNT molecules take U-shape, and the polar headgroups of such molecules cover the pore's interior.

Interaction of ganglioside-containing planar bilayers with serotonin and inorganic cations.

The binding of serotonin and inorganic cations K+, Na+, Ca2+, Mg2+ to planar bilayers formed from mixtures of phosphatidylcholine and mono-, di- and trisialogangliosides was studied by the potentiodynamic and nonactin-induced potassium conductivity method. The theoretical analysis of the results obtained was made taking into account (1) protrusion of the ganglioside charges from the membrane surface and (2) simultaneous adsorption of ions on the bilayer surface and on the ganglioside charges protruding into the solution. It was shown that there was no specific binding of K+ and Na+. The binding constants for Ca2+, Mg2+ were determined. These constants for all the gangliosides studied were equal to 500 M-1. The determined binding constants of serotonin to various gangliosides diminish in the following order: GD3 greater than GT1b greater than GD1a greater than GM1.

[Neutral glycosphingolipids in Ehrlich ascites carcinoma cells]. / Neitral'nye glikosfingolipidy kletokk astsitnoi kartsinomy Erlikha.

The structure of the neutral glycosphingolipids of the Ehrlich ascite carcinoma (EAC) cells was studied. The main four components were identified as glycosylceramide, lastosylceramide, N-acetylgalactosyllactosylceramide and galactosyl-N-acetyllactosylceramide (asialo-GM1). The neutral glycolipid pattern of the cells was found to depend on their density. Dilution of the cell suspension resulted in an increased content of asia-lo-GM1, whereas the content of the other neutral glycolipids remained unchanged. The possible connection between these changes and the earlier disclosed cell density dependence of the gangliosides in EAC cells is discussed.

Changes of glycolipids dependent on cell density of Ehrlich ascites carcinoma cells.

The glycolipid composition of Ehrlich ascites carcinoma cells was found to depend strongly on the cell density of the suspension. The general trend observed upon dilution of the cell suspension was a reduction of the less complex gangliosides GM3 and GM2 with concomitant increase of the more complex gangliosides, especially GM1. The increase of the content of ganglioside GM1 upon dilution was accompanied by a comparable decrease of the content of its immediate precursor, asialo-GM1, whereas the content of other neutral glycosphingolipids did not change very much. When the cell suspension was diluted with medium conditioned by dense cells the ganglioside profile of the diluted suspension remained similar to that of the dense cell suspension. It is postulated that the medium conditioned with dense cells contains a transferable factor inhibiting sialylation of asialo-GM1.

[Relationship between the ganglioside composition of Ehrlich ascitic carcinoma and cell population density]. / Zavisimost' gangliozidnogo sostava astsitnoi kartsinomy Erlikha ot plotnosti kletochnoi populiatsii.

The ganglioside composition of Ehrlich ascite carcinoma cells changes drastically with changes in density of the cell population. Upon 30-fold dilution of the native cell suspension and subsequent incubation for 2 h the amount of total gangliosides per cell doubles the quantities of a monosialoganglioside and of a disialoganglioside increase ten- and five-fold, respectively, whereas the other cell gangliosides do not change. The decrease of cell density is supposed to be accompanied by induction on the cell surface of a sialytransferase activity, which appears to be depressed in the dense suspension.

Unusual gangliosides of eggs and embryos of the sea urchin Strongylocentrotus intermedius. Structure and density-dependence of surface localization.

From eggs and embryos of the sea urchin Strongylocentrotus intermedius two gangliosides, provisionally named G-1 and G-2, were isolated in the pure state. Both gangliosides contained glucose, N-glycoloylneuraminic acid and sphingosines in a 2:2:1 ratio; G-2 contained also a sulfate group, and yielded G-1 on desulfation. By periodate oxidation/borohydride reduction, permethylation analysis, neuraminidase degradation, analysis of the aldohexitol acetates and mass-spectrometry G-1 and G-2 were shown to have hitherto unknown structures: G-1 was identified as N-glycoloylneuraminosyl-(alpha 2 leads to 6)-glucosyl-(1 leads to 8)-N-glycoloylneuraminosyl-(2 leads to 6)-glucosyl-(1 leads to 1)-ceramide, and G-2 as sulfated G-1, carrying a sulfate ester group at C-8 of the terminal sialic acid. Antisera against the two gangliosides were prepared in rabbits by immunization with ganglioside G-1 or G-2. The specificity of the antisera was revealed by immunoelectrophoresis and immunodiffusion. The antisera did not react with bovine-brain and rat-liver gangliosides, with glucosylceramide and with various hydrolytic fragments of G-1 and G-2. The surface localization of the gangliosides in embryos incubated at different cell densities was studied by immunofluorescence microscopy. The intensity of the immunofluorescence was found to increase with decreasing cell density, indicating a different surface organization in sparse and dense embryos. In the sparse embryos immunofluorescence was seen mainly in the contact regions between the blastomers.

Role of gangliosides in reception of influenza virus.

The ganglioside composition of Ehrlich ascites carcinoma (EAC) cells and the role of the individual gangliosides in binding and penetration into the cell of influenza virus were determined. EAC gangliosides identical with or close to GM3, GM2, GM1, GT1a and GT1b were characterized by thin-layer chromarography, compositional analyses, methylation analysis and mass-spectrometry. The ganglioside uptake capacity of native and neuraminidase-treated EAC cells was studied with tritium-labeled gangliosides of definite structure and the binding of influenza virus to cells was determinated by using [3H]uridine-labeled virus and by hemagglutination studies. Treatment of the cells with Vibrio cholerae neuraminidase largely decreased binding of the virus. Exogenous gangliosides with a terminal galactose unit or a penultimate galactose masked by neuraminic acid were able to restore the virus-binding capacity of neuraminidase-treated cells, however, the main ganglioside of EAC cells, GM2, which carbohydrate chain is terminated by N-acetylgalactosamine, was completely ineffective. The common carbohydrate sequence of the gangliosides showing binding activity (formula; see text) is proposed to be the main recognition structure of the influenza virus receptor on the surface of EAC cells. Penetration of labeled influenza virus into the nuclei of EAC cells was evaluated by measuring the radioactivity of the nuclei of neuraminidase-treated ganglioside-loaded cells after exposition to the labeled virus. Of all gangliosides tested only trisialogangliosides of the GT1b type were able to induce increased entry of the virus into the cells and accumulation of its radioactive component into the nuclei. It is suggested that GT1b gangliosides react specifically with the virus protein responsible for membrane fusion (apparently the hemagglutinin HA2 subunit) and thus are involved in virus penetration and delivery of the virus genome to the nuclei.

Immunochemical study of gangliosides at the cell surface of sea urchin embryos.

Two main gangliosides (G-1 and G-2) were isolated from eggs and embryos S. intermedius. They contain glucose, N-glucolylneuraminic acids, phytosphyngosine, fatty acids and alpha-hydroxy fatty acids. Molar ratios and sequence of these components are the same for both gangliosides, but G-2 contains sulphate residue which is attached to the terminal neuraminic acid. To obtain specific antisera rabbits were immunized by G-1 or G-2, which were mixed with bovine serum albumin and Freund's adjuvant. Both gangliosides possessed electrophoretic and antigenic heterogeneity. G-1 and G-2 gangliosides have common and individual antigenic determinants. Glucosylceramide of gangliosides is immunologically inactive. Individual antigenic specificity of the gangliosides depends on the presence of N-glycolylneuraminic acid (G-1) and SO3H-group (G-2). Egg gangliosides were demonstrated by immunofluorescence throughout the cell surface. After fertilization of immunofluorescent label was concentrated on one pole of the embryo only. During the development of specific fluorescence was again uniformly distributed at the blastomer surface. The most intense fluorescence was observed in the junction areas of the blastomers.