Showing papers on "Membrane lipids published in 1984"

Spectrophotometric detection of lipid conjugated dienes.

Abstract: Publisher Summary Like many other substances, naturally occurring lipids exhibit simple end absorption in ultraviolet light as the wavelength is lowered toward 200 nm. The spectra of a variety of organic molecules containing conjugated dienes, however, are characterized by intense absorption, the so-called K band, which may range, with respect to peak absorption, from 215 to 250 nm, depending on nearby substituent groups. Ultraviolet spectrophometric detection of conjugated dienes has been used for many years in the food industry for the detection of autoxidized lipids. The method appears to have been applied for the first time to the problem of liver cell lipid peroxidation of toxigenic origin in 1966 and has been widely used since. For a variety of pathological processes, the question has been raised whether peroxidative decomposition of membrane lipids has occurred in vivo. A second principle of the method recognizes that for whole-animal studies involving possible lipid peroxidation, the fraction of endogenous lipids actually peroxidized may not only be low, but the process of lipid peroxidation may be confined to a particular subcellular structure.

Role of cholesterol in fusion of Semliki Forest virus with membranes.

Abstract: The low pH-triggered membrane fusion activity of Semliki Forest virus is dependent on the presence of cholesterol in the target membrane. When liposomes containing phospholipids and cholesterol analogs were used, fusion activity was observed with steroids which did not have a planar nucleus or an isooctyl side chain at C-17, but fusion activity was not observed when analogs which lacked the 3 beta-OH group were used. Binding of virus to liposomes at low pH was similarly, but not totally, dependent on the presence of a 3 beta-OH sterol.

Physical properties and lipid composition of brain membranes from ethanol tolerant-dependent mice

Abstract: DBA/2 mice were made tolerant to and dependent on ethanol by administration of an ethanol-containing liquid diet for 7 days. Fluorescent probe molecules were used to estimate the fluidity and ethanol sensitivity of brain synaptic membranes from these mice. The fluorescence polarization of cis- parinarate , trans- parinarate , and 1,6-diphenyl-1,3,5-hexatriene (probes of the membrane core) and 1-(4- trimethylammoniumphenyl )-6-phenyl-1,3,5-hexatriene (a probe of the membrane surface) was higher in membranes from ethanol tolerant-dependent mice than in membranes from control mice. The decrease in fluorescence polarization produced by in vitro exposure to ethanol was attenuated in membranes from ethanol tolerant-dependent mice when 1,6-diphenyl-1,3,5-hexatriene was used as the probe, but not when 1-(4- trimethylammoniumphenyl )-6-phenyl-1,3,5-hexatriene was used. These results indicate that chronic ingestion of ethanol decreased the fluidity and the ethanol sensitivity of the synaptic membranes. In contrast to the alterations observed with intact membranes, liposomes of lipids extracted from synaptic membranes of control and ethanol tolerant-dependent mice did not differ in their physical properties. Analysis of membrane lipids demonstrated that chronic ethanol treatment selectively decreased the unsaturated acyl groups of phosphatidylserine without altering the acyl composition of other phospholipids or sphingolipids. The amount of each phospholipid was not changed, but membrane cholesterol was decreased by chronic ethanol ingestion. Use of 2-dimensional thin-layer chromatography allowed the quantitation of 10 different gangliosides. The concentrations of these lipids were unchanged in synaptic membranes from ethanol tolerant-dependent mice. Thus, the changes in membrane physical properties produced by chronic ingestion of ethanol may be due, at least in part, to altered acyl composition of phosphatidylserine. The differences observed between intact membranes and extracted lipids suggest, however, that chronic ethanol treatment also produced changes in the lipid arrangement or lipid-protein interactions of the intact membranes.

Dynamic structure of membranes by deuterium NMR

Abstract: Progress in our understanding of the dynamic structure of membrane lipids and proteins has recently been made possible by the advent of high-field "solid-state" nuclear magnetic resonance spectroscopic studies of specifically deuterium-labeled systems. Major features of lipid and protein dynamics have been deduced.

Diffusion and regionalization in membranes of maturing ram spermatozoa.

Abstract: An essential feature of the "fluid mosaic model" (Singer, S. J., and G. L. Nicolson , 1972, Science (Wash. DC)., 175:720-731) of the cell plasma membrane is the ability of membrane lipids and proteins to diffuse laterally in the plane of the membrane. Mammalian sperm are capable of overcoming free random diffusion and restricting specific membrane components, both lipid and protein, to defined regions of the sperm's surface. The patterns of these regionalizations evolve with the processes of sperm differentiation: spermatogenesis, epididymal maturation, and capacitation. We have used the technique of fluorescence recovery after photobleaching to measure the diffusion of the lipid analogue 1,1'- dihexadecyl 3,3,3',3'- tetramethylindocarbocyanine perchlorate ( C16dil ) on the different morphological regions of testicular and ejaculated ram spermatozoa. We have found: (a) that the major morphologically distinct regions (head, midpiece, and tail) of the plasma membrane of both testicular and ejaculated spermatozoa are also physically distinct as measured by C16dil diffusibility; (b) that despite regional differences in diffusibility there is exchange of this lipid analogue by lateral diffusion between the major morphological regions of the plasma membrane; and (c) that epididymal maturation results in changes in C16dil diffusibility in the different regions of the sperm plasma membrane. In particular, the plasma membranes of the anterior and posterior heads become physically distinct.

Detection of erythrocyte membrane proteins, sialoglycoproteins, and lipids in the same polyacrylamide gel using a double-staining technique

Abstract: A silver/Coomassie brilliant blue R-250 staining technique that permits a color-coded differentiation of erythrocyte membrane proteins, sialoglycoproteins, and lipids in a single one-dimensional NaDodSO4/polyacrylamide gel has been described. Gels stained first with silver stain and then with Coomassie blue (CB) showed the characteristic blue staining of all conventional CB-sensitive membrane polypeptides, whereas periodic acid-Schiff reagent-sensitive sialoglycoproteins and lipids stained yellow. Several yellow Ag-stained bands corresponding to major and minor sialoglycoproteins were detected at Mr X 10(-3) of 88, 72, 65, 41, 35, 31, 28, 24, and 20. Neuraminidase treatment of intact erythrocytes caused shifts in the electrophoretic mobilities of several yellow-stained bands without affecting the CB-stained polypeptide pattern. These observations afforded evidence that the yellow-staining bands were sialoglycoproteins and lipids. The double-staining technique was used in a topological analysis of the membrane surface of the erythrocyte using protease digestion and selective solubilization. Trypsin cleaved the yellow bands at Mr 88,000 and 41,000. Membrane-associated cleavage products were noted at Mr 58,000 and 38,000. Pronase treatment of intact cells gave membrane-associated cleavage products at Mr 38,000 (yellow) and two CB-stained bands at Mr 58,000 and 60,000. These results suggested that the double-staining technique may be applicable in compositional and topological analyses of other biological membranes.

Voltage-Regulated Sodium Channel Molecules

Abstract: In summary, TTX and STX binding have been used to follow the purification of sodium channel proteins from electric organ, mammalian skeletal muscle, and brain. In each instance they were proteins somewhat larger than the acetylcholine receptor and exhibited stability properties that reflect an intimate interaction with membrane lipids. The principal peptide constituent seems to be a large glycopeptide of approximately 250,000 daltons. Because this is evidently the only constituent of the electroplax protein, it must contain the TTX receptor site, probably forming the ion pathway itself. Photo-labeling with ScTX and studies with Tityus gamma toxin (58a) indicate a site involved with gating processes, also associated with the large peptide of the neuronal and electroplax proteins. The smaller peptides are not consistent features of all of the preparations, but may contribute to the molecular ensemble. If, however, the entire channel were formed from a single extremely large peptide, there would be interesting mechanistic implications, because the ion transporting and voltage-sensing mechanisms would be accounted for by domains within the folded polypeptide chain. Clearly, the prospect for combining biochemical isolation and reconstitution with the new biophysical technology offers an exciting experimental conjunction.

Chloroplast Phospholipid Molecular Species Alterations during Low Temperature Acclimation in Dunaliella

Abstract: A detailed analysis of the low temperature-induced alterations of Dunaliella salina (UTEX 1644) microsomal membrane lipids was carried out. Microsomal membranes were isolated from cells grown at 30°C, from cells shifted to 12°C for 12 hours, and from cells acclimated to 12°C. Fatty acid analyses of the major lipid classes demonstrated significant changes in the fatty acid composition of phosphatidylcholinemine (PE) and phosphatidylglycerol (PG) but not phosphatidylcholine (PC) during the initial 12 hours at low temperature. These changes did not entail enhanced desaturation of linoleic acid. Subsequent to 12 hours, the proportions of linolenic acid increased in all phospholipids. Molecular species analyses of the phospholipids demonstrated that the most immediate changes following a shift to low temperature were limited to several molecular species of PE and PG. The changes observed in PE included a decrease in C 30 species and concomitant increases in C 34 and C 36 species. Compositional changes associated with PG entailed the emergence of a new molecular species (18:1/18:1) not found at 30°C. The retailoring of molecular species resulted in an increase in the number of species having two unsaturated acyl chains and did not reflect a simple enhancement of desaturase activity as suggested by the fatty acid analysis. We conclude that the initial alterations in response to low temperature stress involve discrete changes in certain molecular species. These and further alterations of molecular species following acclimation to low temperature would appear to augment increases in acyl chain desaturation as a means of modifying membrane properties in response to low temperature stress.

Lipid Bilayer Stability in Biological Membranes

Abstract: One of the most important problems in biophysics today is the self-assembly of membrane components, in particular lipids and proteins. It has long been known that many membrane lipids spontaneously form a bilayer when mixed with water (Luzzati, 1968). Recently, it was also indicated that even a membrane protein may form a bilayer structure (Carlsson, 1981). The Singer and Nicolson (1972) model of a biomembrane is based on the assumption that the lipids form a bilayer matrix in which the proteins are incorporated and are able to diffuse more or less freely in two dimensions. If the function of the lipids is only to form this fluid matrix, why does a biological membrane often contain more than 100 different lipid species? Furthermore, many lipids do not spontaneously form bilayers with water. Sometimes not even the major lipid in a membrane forms a bilayer, e.g., monogalactosyldiglyceride of chloroplasts (Shipley et al., 1973; Brentel et al., 1984c). However, the membrane lipids together with the proteins form a stable, functioning membrane, with the properties necessary for a living cell, i.e., to be both a barrier and a communicator to the surroundings. It can thus be expected that the membrane lipids are not working just as a fluid matrix, as is suggested by the Singer and Nicolson model. Most probably they play an important structural role in biological membranes as will be discussed in this chapter.

Role of membrane lipids in peptide hormone function: binding of enkephalins to micelles

Abstract: In the course of their biological function, peptide hormones must be transferred from an aqueous phase to the lipid-rich environment of their membrane-bound receptor proteins. We have investigated the possible influence of phospholipids in this process, using 360-MHz 1H and 90-MHz 13C NMR spectroscopy to examine the association of the opioid peptides [Met]- and [Leu]enkephalins (Tyr-Gly-Gly-Phe-Met/Leu) with phospholipid micelles. Binding of peptides to lipid was monitored in NMR spectra by selective chemical shift movements (e.g., the Phe aromatic ring protons) and residue-specific line broadening (e.g., of Met/Leu carbonyl- and alpha-carbon resonances). Results established that the zwitterionic hormones associate hydrophobically both with a neutral lipid (lysophosphatidylcholine) and (also electrostatically) with a negative lipid (lysophosphatidylglycerol). An association constant of Ka = 3.7 X 10(1) M-1 was calculated for the hydrophobic binding of enkephalin to lysophosphatidylcholine. NMR data suggested that enkephalin binds to the lipid with Met/Leu, Phe, and likely Tyr side-chain substituents associated with nonpolar interior regions of the micelle, whereas the COOH-terminal carboxylate moiety of the peptide is located in the surface of the lipid particle. An "attraction-interaction" model is proposed for hormone-lipid association wherein negative lipids attract the hormone electrostatically, while site-specific hydrophobic contacts facilitate its entry, concentration, and orientation into the lipid phase.

Regulation of Membrane Fluidity by Lipid Desaturases

Abstract: It has long been known that normal prokaryotic as well as eukaryotic cells can grow only when their membrane lipids are largely in the fluid state, i.e., at temperatures above the gel to liquid-crystalline transition temperature (T m) of their membrane lipids (see McElhaney, this volume). Adaptation of bacteria (Cronan, 1975; Fulco, this volume), yeast (Watson, this volume), fungi (Miller and Barran, this volume), higher plants (Mazliak, 1979), and the protozoan Tetrahymena (Thompson and Nozawa, this volume) to temperatures below their normal growth temperatures generally results in changes in membrane lipid composition leading to increases in fatty acid unsaturation. The major factor affecting the fluidity of membrane lipids in eukaryotes, apart from the presence of cholesterol, is the degree of unsaturation of their fatty acid chains. This holds also for prokaryotes but, in addition, other factors such as chain length and branching may be important.

Homeostatic control of membrane fatty acid composition in the rat after dietary lipid treatment

Abstract: Diets in which both the lipid content and composition (polyunsaturated to saturated fatty acid ratio) were varied were fed to rats for 20 weeks, and the effects on the tissue lipid profiles were determined. The fatty acid profile of the plasma lipids, and the phospholipid fatty acids of the mitochondrial and microsomal fractions of liver, heart, kidney and brain, as well as erythrocyte membranes were determined. Despite large differences in the level and type of lipid present in the experimental diets and in the proportion of saturated fatty acids in the plasma lipids in response to the various diets, there was little effect on the proportion of saturated to unsaturated fatty acids in the phospholipids of the various membranes examined. The major effect of altering the dietary level of polyunsaturated to saturated fatty acids was on the ratio of the omega 6/omega 3 series of unsaturated fatty acids in the membrane lipids. This change occurred in all tissues except the brain, in which only a small response to altered dietary lipid intake was observed. The omega 6/omega 3 ratio was elevated upon feeding a diet rich in omega 6 polyunsaturated fatty acids, but decreased when a diet rich in saturated fatty acids was fed. The failure to significantly alter membrane lipid saturation/unsaturation in the tissues examined would suggest that a homeostatic mechanism is operative in biological membranes and may act to buffer membranes from the effects of changes in the nature of the dietary lipid intake.

Lateral mobility of phospholipid and cholesterol in the human erythrocyte membrane: effects of protein-lipid interactions.

Abstract: The phospholipid and cholesterol derivatives N-(7-nitro-2,1,3-benzoxadiazol-4-yl)phosphatidylethanolamine (NBD-PE) and N1-cholesterylcarbamoyl-N8-(7-nitro-2,1,3-benzoxadiazol-4-yl )-3,6-dioxaoctane-1 , 8-diamine (NBD-Chol), respectively, were incorporated into egg phosphatidylcholine/cholesterol multilamellar liposomes, human erythrocyte ghost membranes, and multilamellar liposomes derived from extracted human erythrocyte membrane lipids. The lateral mobility of these probes in the plane of the various membranes was measured by using the fluorescence photo-bleaching recovery technique. NBD-PE and NBD-Chol manifested identical lateral mobilities in egg phosphatidylcholine/cholesterol multilamellar liposomes over the range of temperatures from 10 to 37 degrees C and the range of cholesterol mole fractions from 0.0 to 0.5, and in erythrocyte ghost membranes and erythrocyte membrane lipid-derived multilamellar liposomes over the range of temperatures from 15 to 37 degrees C. The weak temperature dependence of the lateral diffusion coefficients of the lipid probes in both artificial and erythrocyte ghost membranes is consistent with the lack of a phase transition in any of these systems over the temperature range studied. Both NBD-PE and NBD-Chol diffuse 4-fold faster in liposomes derived from extracted erythrocyte membrane lipids (D = 8.0 X 10(-9) cm2 s-1 at 37 degrees C) than in the ghost membranes themselves (D = 2.1 X 10(-9) cm2 s-1 at 37 degrees C), suggesting a significant restriction of lipid lateral mobility by membrane protein in the human erythrocyte membrane.

Lipid Composition of a Psychrophilic Marine Vibrio sp. During Starvation-Induced Morphogenesis.

Abstract: Qualitative and quantitative changes with time in phospholipids and fatty acids were examined after suspension of cells of a psychrophilic marine bacterium in nutrient-free artificial seawater at 5°C. Viability was maintained throughout the 21-day examination period, with plate counts and acridine orange direct counts indicating a slight increase in cell number. Gravimetric data, however, showed a significant decrease in bacterial biomass during the 3-week study. Levels of ATP per cell also decreased significantly (59%) during the starvation period. Since starvation (resulting in dormancy) is probably the typical physiological state of marine bacteria, estimation of bacterial density in marine waters by using ATP data obtained from log-phase cells is probably inaccurate. Total lipid phosphate decreased (65%) during the starvation period, with phosphatidylethanolamine showing the greatest loss. A large increase (57%) in the neutral lipid fraction was also detected, especially during the first week of starvation. A selective increase in palmitoleate at the expense of myristate was detected in the membrane lipids. The effects of these changes on membrane fluidity and the possible consequences for these cells in the marine environment are discussed.

Membrane Lipids and Enzymes of Cultured High- and Low-Metastatic B16 Melanoma Variants

Abstract: B16 melanoma cell variants were used to determine if the metastatic properties of these cells could be correlated to distinct plasma membrane, microsome, and mitochondrial membrane lipid compositions and membrane-bound enzyme activities in high- and low-metastatic cell variants, respectively. The high-metastatic B16-F10 melanoma cell membranes had lower cholesterol/phospholipid ratios, lower arachidonic acid content, lower polyunsaturated fatty acid content, higher phosphatidylcholine/phosphatidylethanolamine ratios, and higher succinate cytochrome c reductase activity than those of B16-F1 melanoma cell membranes. No differences in cholesterol/phospholipid ratio were noted in the mitochondria. Na+-K+-adenosinetriphosphatase activity and solubility of 5'-nucleotidase activity were also similar. The data indicate that the membrane lipid composition of B16-F10 melanoma cells is distinct from that of B16-F1 melanoma cells and may help to elucidate the molecular basis for the different metastatic properties of these cell lines in vivo.

Outer-membrane protein PhoE from Escherichia coli forms anion-selective pores in lipid-bilayer membranes.

Abstract: Porin PhoE of the outer membrane of Escherichia coli was isolated and purified. Reconstitution experiments with lipid bilayer membranes showed that this protein formed pores which had a single channel conductance of 210 pS at 0.1 M KCl. The PhoE pores were obviously not voltage-controlled or regulated. In contrast to pores formed by the OmpF porin from E. coli the PhoE channel was found to be anion-selective at neutral pH. Chloride is about three to ten times more permeable through the pore than alkali ions. On the basis of the observed pH dependence of the permeability ratio of anions and cations, this anionic selectivity is explained by the assumption that the PhoE pore contains an excess of fixed positive charges.

Assignment of acyl chain resonances from membranes of mammalian cells by two-dimensional NMR methods

Abstract: Two-dimensional nuclear magnetic resonance (NMR) methods have been successfully used to assign resonances in the 1H NMR spectrum of intact viable rat mammary adenocarcinoma cells. Two-dimensional scalar-correlated spectroscopy identifies connectivities for resonances of the lipid acyl chains in the plasma membrane of these cells. We expect that two-dimensional scalar-correlated methods may be of general use for providing unequivocal assignments in the complex and often poorly resolved 1H NMR spectra of cells.

Isolation and characterization of the Legionella pneumophila outer membrane.

Abstract: A whole cell lysate of Legionella pneumophila was fractionated into five membrane fractions by sucrose gradient centrifugation. Membranes were characterized by enzymatic, chemical, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. Two forms of cytoplasmic membrane (CM-1, CM-2), a band of intermediate density (IM), and two forms of outer membrane (OM-1, OM-2) were detected. The CM-1 fraction was the purest form of cytoplasmic membrane, and fraction CM-2 was primarily cytoplasmic membrane associated with small amounts of peptidoglycan. The IM, CM-1, and CM-2 fractions were enriched in peptidoglycan, and the amount of carbohydrate and 2-keto-3-deoxyoctonic acid was not appreciably greater in outer membrane relative to cytoplasmic membrane. Phosphatidylethanolamine and phosphatidylcholine were found to be the major phospholipids in the membrane fractions. The major outer membrane proteins had molecular sizes of 29,000 and 33,000 daltons and were both modified by heating. The 29,000-dalton protein was tightly associated with the peptidoglycan and was equally distributed in the IM, OM-1, and OM-2.